Prodrugs of thrombin inhibitors

ABSTRACT

There is provided compounds of formula I,
 
R 1 O(O)C—CH 2 —(R)Cgl-Aze-Pab-R 2 I
 
wherein R 1  and R 2  have meanings given in the description, which are useful as prodrugs of inhibitors of trypsin-like proteases, such as thrombin, and in particular in the treatment of conditions where inhibition of thrombin is required (eg thrombosis) or as anticoagulants.

This application is a continuation of U.S. application Ser. No.10/074,008, filed Feb. 14, 2002, now abandoned, which is a continuationof U.S. application Ser. No. 09/708,449, filed Nov. 9, 2000, nowabandoned, which is a continuation of U.S. application Ser. No.09/353,644, filed Jul. 15, 1999, now U.S. Pat. No. 6,262,028, which is acontinuation of U.S. application Ser. No. 08/776,231, filed Jan. 31,1997, now U.S. Pat. No. 5,965,692, which is a 371 of PCT/SE96/01680,filed Dec. 17, 1996, the entire content of which is hereby incorporatedby reference in this application.

FIELD OF THE INVENTION

This invention relates to pharmaceutically useful prodrugs ofpharmaceutically active compounds, which active compounds are, inparticular, competitive inhibitors of trypsin-like serine proteases,especially thrombin, the use of the prodrugs as medicaments,pharmaceutical compositions containing them and synthetic routes totheir production.

BACKGROUND

Blood coagulation is the key process involved in both haemostasis (iethe prevention of blood loss from a damaged vessel) and thrombosis (iethe formation of a blood clot in a blood vessel, sometimes leading tovessel obstruction).

Coagulation is the result of a complex series of enzymatic reactions.One of the ultimate steps in this series of reactions is the conversionof the proenzyme prothrombin to the active enzyme thrombin.

Thrombin is known to play a central role in coagulation. It activatesplatelets, leading to platelet aggregation, converts fibrinogen intofibrin monomers, which polymerise spontaneously into fibrin polymers,and activates factor XIII, which in turn crosslinks the polymers to forminsoluble fibrin. Furthermore, thrombin activates factor V and factorVIII leading to a “positive feedback” generation of thrombin fromprothrombin.

By inhibiting the aggregation of platelets and the formation andcrosslinking of fibrin, effective inhibitors of thrombin would thereforebe expected to exhibit antithrombotic activity. In addition,antithrombotic activity would be expected to be enhanced by effectiveinhibition of the positive feedback mechanism.

PRIOR ART

The development of low molecular weight inhibitors of thrombin has beendescribed by Claesson in Blood Coagul. Fibrin. (1994) 5, 411.

Blombāck et al (in J. Clin. Lab. Invest. 24, suppl. 107, 59, (1969))reported thrombin inhibitors based on the amino acid sequence situatedaround the cleavage site for the fibrinogen Aα chain. Of the amino acidsequences discussed, these authors suggested the tripeptide sequencePhe-Val-Arg would be the most effective inhibitor.

Low molecular weight peptide-based thrombin inhibitors have subsequentlybeen disclosed in, for example, U.S. Pat. No. 4,346,078; InternationalPatent Applications WO 93/11152, WO 94/29336, WO 93/18060 and WO95/01168; and European Patent Applications 648 780, 468 231, 559 046,641 779, 185 390, 526 877, 542 525, 195 212, 362 002, 364 344, 530 167,293 881, 686 642 and 601 459.

More recently, thrombin inhibitors based on peptide derivatives havebeen disclosed in European Patent Application 0 669 317 andInternational Patent Applications WO 95/23609, WO 95/35309, WO 96/25426and WO 94/29336.

In particular, the latter application discloses the peptide derivativesR^(a)OOC—CH₂—(R)Cgl-Aze-Pab-H, wherein R^(a) represents H, benzyl orC₁₋₆ alkyl.

Although these active compounds are known to exhibit significantantithrombin activity, it would be beneficial to improve theirpharmacokinetic properties both after oral and parenteraladministration. Examples of pharmacokinetic properties which it isdesirable to improve include:

-   -   (a) providing an improved absorption from the gastrointestinal        tract, with a view to reducing intra- and/or inter-individual        variability in relation to the bioavailability of the active        compounds;    -   (b) flattening the plasma concentration time profile (ie        reducing the peak/trough ratio in the plasma concentration over        the dosing interval), with a view to reducing the risk of        falling outside the therapeutic interval and the side effects        caused by a concentration peak which is too high (eg bleeding),        and those caused by one which is too low (eg thrombus        formation); and    -   (c) increasing the duration of action of the active compounds.

Moreover, oral and parenteral administration of active thrombininhibitors may lead to undesirable local bleeding (eg in the intestinallumen or subcutaneously) as a result of a high local concentration,

Finally, orally administered active thrombin inhibitors which alsoinhibit trypsin and other serine proteases in the gastrointestinal tractmay exhibit additional side effects, including indigestion (eg iftrypsin is inhibited in the intestinal lumen).

Although certain N-benzyloxycarbonyl derivatives of the aforementionedactive compounds are also disclosed as thrombin inhibitors inInternational Patent Application WO 94/29336, that these derivatives maybe useful as prodrugs is not mentioned. In fact, WO 94/29336 makes nomention of suitable prodrugs of the active compounds.

We have found that the above problems may be solved by administeringcompounds according to the present invention which, whilst inactive perse, upon oral and/or parenteral administration are metabolised in thebody to form active thrombin inhibitors, including those mentionedabove.

DISCLOSURE OF THE INVENTION

According to the invention there is provided a compound of formula I,R¹O(O)C—CH₂—(R)Cgl-Aze-Pab-R²  Iwherein

-   R¹ represents —R³ or -A¹C(O)N(R⁴)R⁵ or -A¹C(O)OR⁴;-   A¹ represents C₁₋₅ alkylene;-   R² (which replaces one of the hydrogen atoms in the amidino unit of    Pab-H) represents OH, OC(O)R⁶, C(O)OR⁷ or C(O)OCH((R⁸))OC(O)R⁹;-   R³ represents H, C₁₋₁₀ alkyl, or C₁₋₃ alkyphenyl (which latter group    is optionally substituted by C₁₋₆ alkyl, C₁₋₆ alkoxy, nitro or    halogen);-   R⁴ and R⁵ independently represent H, C₁₋₆ alkyl, phenyl, 2-naphthyl    or, when R¹ represents -A¹C(O)N(R⁴)R⁵, together with the nitrogen    atom to which they are attached represent pyrrolidinyl or    piperidinyl;-   R⁶ represents C₁₋₁₇ alkyl, phenyl or 2-naphthyl (all of which are    optionally substituted by C₁₋₆ alkyl or halogen);-   R⁷ represents 2-naphthyl, phenyl, C₁₋₃ alkylphenyl (which latter    three groups are optionally substituted by C₁₋₆ alkyl, C₁₋₆ alkoxy,    nitro or halogen), or C₁₋₁₂ alkyl (which latter group is optionally    substituted by C₁₋₆ alkoxy, C₁₋₆ acyloxy or halogen);-   R⁸ represents H or C₁₋₄ alkyl; and-   R⁹ represents 2-naphthyl, phenyl, C₁₋₆ alkoxy or C₁₋₈ alkyl (which    latter group is optionally substituted by halogen, C₁₋₆ alkoxy or    C₁₋₆ acyloxy); provided that when R¹ represents R³, R³ represents    benzyl, methyl, ethyl, n-butyl or n-hexyl and R² represents C(O)OR⁷,    then R⁷ does not represent benzyl;-   or a pharmaceutically-acceptable salt thereof (hereinafter referred    to as “the compounds of the invention”).

The compounds of the invention may exhibit tautomerism. All tautomericforms and mixtures thereof are included within the scope of theinvention.

The compounds of the invention may also contain one or more asymmetricis carbon atoms and may therefore exhibit optical and/ordiastereoisomerism. All diastereoisomers may be separated usingconventional techniques, eg chromatography or fractionalcrystallisation. The various stereoisomers may be isolated by separationof a racemic or other mixture of the compounds using conventional, egfractional crystalaisation or HPLC, techniques. Alternatively thedesired optical isomers may be made by reaction of the appropriateoptically active starting materials under conditions which will notcause racemisation or epimerisation, or by derivatisation, for examplewith a homochiral acid followed by separation of the diastereomericderivatives by conventional means (eg HPLC, chromatography over silica).All stereoisomers are included within the scope of the invention.

According to a further aspect of the invention there is provided the useof a compound of formula I, as hereinbefore defined but without theproviso, as a prodrug.

Alkyl groups which R³, R⁴, R⁵, R⁶, R⁷ and R⁹ may represent may be linearor, when there are a sufficient number of carbon atoms, be branched, becyclic or partially cyclic, be saturated or unsaturated, be interruptedby oxygen and/or be substituted or terminated by OH, provided that theOH group is not attached to an sp² carbon atom or a carbon atom which isadjacent to an oxygen atom.

By “partially cyclic alkyl groups” we mean groups such as CH₂Ch.

Alkyl groups which R⁸ may represent, and R³, R⁶ and R⁷ may besubstituted by, may be linear or, when there are a sufficient number ofcarbon atoms, be branched, be saturated or unsaturated and/or beinterrupted by oxygen.

The alkyl portion of alkylphenyl groups which R³ and R⁷ may representmay be linear or, when there are a sufficient number of carbon atoms, bebranched and/or be saturated or unsaturated.

Alkylene groups which A¹ may represent may be linear or, when there area sufficient number of carbon atoms, be branched and/or be saturated orunsaturated.

Alkoxy groups which R⁹ may represent, and R³, R⁷ and R⁹ may besubstituted by, may be linear or, when there are a sufficient number ofcarbon atoms, be branched and/or be saturated or unsaturated.

Acyloxy groups which R⁷ and R⁹ may be substituted by may be linear or,when there are a sufficient number of carbon atoms, be branched and/orbe saturated or unsaturated.

Abbreviations are listed at the end of this specification.

According to a further aspect of the invention there is provided acompound of formula I, as hereinbefore defined, with the additionalprovisos that:

-   -   (a) R¹ does not represent -A¹C(O)OR⁴;    -   (b) R⁴ and R⁵ do not independently represent H;    -   (c) R⁶ does not represent C₁₋₁₇ alkyl, when R² represents        OC(O)R⁶.

According to a further aspect of the invention there is provided acompound of formula I, wherein:

-   -   (a) R¹ represents -A¹C(O)OR⁴;    -   (b) R⁴ and R⁵ independently represent H;    -   (c) R⁶ represents C₁₋₁₇ alkyl, when R² represents OC(O)R⁶.

When R¹ represents -A¹C(O)N(R⁴)R⁵, preferred compounds of the inventioninclude those wherein:

-   A¹ represents C₁₋₃ alkylene;-   R⁴ represents H or C₁₋₆ alkyl;-   R⁵ represents C₁₋₆ alkyl or C₄₋₆ cycloalkyl; or those wherein-   R⁴ and R⁵ together represent pyrrolidinyl.

When R¹ represents -A¹C(O)OR⁴, preferred compounds of the inventioninclude those wherein:

-   A¹ represents C₁₋₅ alkylene;-   R⁴ represents C₁₋₆ alkyl.

When R¹ represents R³, preferred compounds of the invention includethose wherein R³ represents H, C₁₋₁₀ alkyl (which latter group may belinear or, when there are a sufficient number of carbon atoms, may bebranched and/or be partially cyclic or cyclic), or C₁₋₃ alkylphenyl(which latter groups is optionally substituted, may be linear or, whenthere are a sufficient number of carbon atoms, be branched).

Preferred compounds of the invention include those wherein R² representsOH, OC(O)R⁶ (wherein, in the latter case, R⁶ represents optionallysubstituted phenyl or C₁₋₁₇ alkyl (which latter group may be linear or,when there are a sufficient number of carbon atoms, may be branched, becyclic or partially cyclic, and/or be saturated or unsaturated)),C(O)OR⁷ (wherein, in the latter case, R⁷ represents optionallysubstituted phenyl, C₁₋₁₂ alkyl (which latter group is optionallysubstituted, may be linear or, when there are a sufficient number ofcarbon atoms, may be branched, be cyclic or partially cyclic, and/orsaturated or unsaturated), or C₁₋₃ alkylphenyl (which latter group isoptionally substituted, may be linear or, when there are a sufficientnumber of carbon atoms, may be branched)), or C(O)OCH(R⁸)OC(O)R⁹(wherein, in the latter case, R⁸ represents H or methyl, and R⁹represents phenyl, or C₁₋₈ alkyl (which latter group is optionallysubstituted, may be linear or, when there are a sufficient number ofcarbon atoms, may be branched and/or cyclic or partially cyclic)).

More preferred compounds of the invention include those wherein: R¹represents H, linear C₁₋₁₀ alkyl, branched C₃₋₁₀ alkyl, partially cyclicC₄₋₁₀ to alkyl, C₄₋₁₀ cycloalkyl, optionally substituted linear C₁₋₃alkylphenyl, optionally substituted branched C₃ alkylphenyl,-A¹C(O)N(R⁴)R⁵ (wherein, in the latter case, A¹ represents C₁₋₃alkylene, and R⁴ represents H or C₁₋₃ alkyl and R⁵ represents C₂₋₆ alkylor C₅₋₆ cycloalkyl, or R⁴ and R⁵ together represent pyrrolidinyl), or-A¹C(O)OR⁴ (wherein, in the latter case, A¹ represents C₁₋₅ alkylene andR⁴ represents C₁₋₄ alkyl); R² represents OH, OC(O)R⁶ (wherein, in thelatter case, R⁶ represents optionally substituted phenyl, linear C₁₋₄alkyl, branched C₃₋₄ alkyl or cisoleyl), C(O)OR⁷ (wherein, in the lattercase, R⁷ represents optionally substituted and/or optionally unsaturatedlinear C₁₋₄ alkyl or optionally substituted and/or optionallyunsaturated branched C₃₋₄ alkyl, optionally substituted phenyl, oroptionally substituted linear C₁₋₃ alkylphenyl or optionally substitutedbranched C₃ alkylphenyl) or C(O)OCH(R⁸)OC(O)R⁹ (wherein, in the lattercase, R⁸ represents H or methyl and R⁹ represents phenyl, C₅₋₇cycloalkyl, linear C₁₋₆ alkyl, branched C₃₋₆ alkyl or partially cyclicC₇₋₈ alkyl).

Particularly preferred compounds of the invention include those wherein:

-   R¹ represents linear C₁₋₆ alkyl, C₆₋₁₀ cycloalkyl, or optionally    substituted linear C₁₋₃ alkylphenyl;-   R² represents OH, OC(O)R⁶ (wherein, in the latter case, R⁶    represents linear C₁₋₃ alkyl or branched C₃ alkyl), C(O)OR⁷    (wherein, in the latter case, R⁷ represents optionally substituted    linear C₁₋₄ alkyl or optionally substituted branched C₃₋₄ alkyl,    optionally substituted linear C₁₋₃ alkylphenyl or branched C₃    alkylphenyl) or C(O)OCH(R⁸)OC(O)R⁹ (wherein, in the latter case, R⁸    represents H and R⁹ represents C₅₋₇ cycloalyl, linear C₁₋₆ alkyl or    partially cyclic C₇₋₈ alkyl).

When R¹ represents R³ and R³ represents optionally substituted C₁₋₃alkylphenyl, preferred optional substituent include C₁₋₆ alkyl(especially methyl).

When R² represents C(O)OR⁷ and R⁷ represents optionally substitutedC₁₋₁₂ alkyl, preferred optional substituents include halogen (especiallychloro) and C₁₋₆ alkoxy (especially methoxy).

When R² represents C(O)OR⁷ and R⁷ represents optionally substitutedphenyl, preferred optional substituents include C₁₋₆ alkyl (especiallymethyl), C₁₋₆ alkoxy (especially methoxy) and halogen (especiallychloro).

When R² represents C(O)OR⁷ and R⁷ represents optionally substituted C₁₋₃alkylphenyl, preferred optional substituents include nitro.

Preferred compounds of the invention include the compounds of Examples 1to 68.

More preferred compounds of the invention include:

-   EtOOCCH₂—(R)Cgl-Aze-Pab-COOCH₂CH═CH₂;-   nPrOOCCH₂—(R)Cgl-Aze-Pab-COOCH₂CH═CH₂;-   tBuOOCCH₂—(R)Cgl-Aze-Pab-COOCH₂CH═CH₂;-   EtOOCCH₂—(R)Cgl-Aze-Pab-COOEt;-   EtOOCCH₂—(R)Cgl-Aze-Pab-COO-nBu;-   PrlC(O)CH₂CH₂CH₂OOCCH₂—(R)Cgl-Aze-Pab-Z;-   ChNHC(O)CH₂OOCCH₂—(R)Cgl-Aze-Pab-Z;-   (nPr)₂NC(O)CH₂OOCCH₂—(R)Cgl-Aze-Pab-COOCH₂OOCC(CH₃)₃;-   EtOOCCH₂—(R)Cgl-Aze-Pab-COOCH₂OOCC(CH₃)₃;-   EtOOCCH₂—(R)Cgl-Aze-Pab-COOCH(CH₃)OOCCH₃;-   MeOOCCH₂—(R)Cgl-Aze-Pab-OOCPh;-   MeOOCCH₂—(R)Cgl-Aze-Pab-OH;-   EtOOCCH₂—(R)Cgl-Aze-Pab-OH;-   BnOOCCH₂—(R)Cgl-Aze-Pab-OH;-   nPrOOCCH₂—(R)Cgl-Aze-Pab-Z;-   nPrOOCCH₂—(R)Cgl-Aze-Pab-OH;-   iPrOOCCH₂—(R)Cgl-Aze-Pab-OH;-   tBuOOCCH₂—(R)Cgl-Aze-Pab-OH;-   (nPr)₂NCOCH₂OOCCH₂—(R)Cgl-Aze-Pab-OH;-   ChNHCOCH₂OOCCH₂—(R)Cgl-Aze-Pab-OH;-   EtOOCCH₂—(R)Cgl-Aze-Pab-OAc;-   HOOCCH₂—(R)Cgl-Aze-Pab-OH;-   HOOCCH₂—(R)Cgl-Aze-Pab-O-cis-Oleyl;-   Cyclooctyl-OOCCH₂—(R)Cgl-Aze-Pab-Z;-   tBuCH₂OOCCH₂—(R)Cgl-Aze-Pab-Z;-   (2-Me)BnOOCCH₂—(R)Cgl-Aze-Pab-Z;-   ChCH₂OOCCH₂—(R)Cgl-Aze-Pab-Z;-   ChOOCCH₂—(R)Cgl-Aze-Pab-Z;-   PhC(Me)₂OOCCH₂—(R)Cgl-Aze-Pab-Z;-   (Me)₂CHC(Me)₂OOCCH₂—(R)Cgl-Az-Pab-Z;-   BnOOCCH₂—(R)Cgl-Aze-Pab-COOPh(4-OMe);-   ChCH₂OOCCH₂—(R)Cgl-Aze-Pab-COOPh(4-OMe);-   (2-Me)BnOOCCH₂—(R)Cgl-Aze-Pab-COOPh(4-OMe);-   EtOOCCH₂—(R)Cgl-Aze-Pab-COOPh(4-Me);-   BnOOCCH₂—(R)Cgl-Aze-Pab-COOPh(4-Me);-   BnOOCCH₂—(R)Cgl-Aze-Pab-COO-nBu;-   iPrOOCCH₂—(R)Cgl-Aze-Pab-COOCH₂CH═CH₂;-   EtOOCCH₂—(R)Cgl-Aze-Pab-COO-iBu;-   BnOOCCH₂—(R)Cgl-Aze-Pab-COO-nPr;-   EtOOCCH₂—(R)Cgl-Aze-Pab-COOCH₂OOCCh;-   EtOOCCH₂—(R)Cgl-Aze-Pab-COOCH₂OOCCH₂Ch;-   EtOOCCH₂—(R)Cgl-Aze-Pab-COOCH(Me)OOCPh;-   EtOOCCH₂—(R)Cgl-Aze-Pab-COOCH₂OOCPh;-   BnOOCCH₂—(R)Cgl-Aze-Pab-COOCH(Me)OAc;-   EtOOCCH₂—(R)Cgl-Aze-Pab-COOCH₂OAc;-   tBuOOCCH₂—(R)Cgl-Aze-Pab-COOCH₂OAc;-   MeOOC—C(═CHEt)CH₂—OOCCH—(R)Cgl-Aze-Pab-Z;-   Men-OOCCH₂—(R)Cgl-Aze-Pab-COOPh(4-OMe); and-   EtOOCCH₂—(R)Cgl-Aze-Pab-COOCH₂CCl₃.

Particularly preferred compounds of the invention include:

-   EtOOCCH₂—(R)Cgl-Aze-Pab-COOCH₂CCl₃;-   BnOOCCH₂—(R)Cgl-Aze-Pab-COOnBu;-   nPrOOCCH₂—(R)Cgl-Aze-Pab-Z;-   Cyclooctyl-OOCCH₂—(R)Cgl-Aze-PabZ;-   EtOOCCH₂—(R)Cgl-Aze-Pab-COOCH₂OOCCh;-   MeOOCCH₂—(R)Cgl-Aze-Pab-OH;-   EtOOCCH₂—(R)Cgl-Aze-Pab-OH;-   nPrOOCCH₂—(R)Cgl-Aze-Pab-OH;-   iPrOOCCH₂—(R)Cgl-Aze-Pab-OH;-   BnOOCCH₂—(R)Cgl-Aze-Pab-OH; and-   EtOOCCH₂—(R)Cgl-Aze-Pab-OAc.    Preparation

According to the invention there is also provided a process for thepreparation of compounds of formula I which comprises:

-   (a) Preparation of a compound of formula I wherein R² represents OH    by reaction of a corresponding compound of formula I, wherein R²    represents OC(O)R⁶ and R⁶ is as hereinbefore defined, with an    alkoxide base (eg an alkali metal alkoxide), for example at room    temperature in the presence of an appropriate organic solvent (eg    THF)-   (b) Preparation of a compound of formula I wherein R² represents OH    by reaction of a corresponding compound of formula I wherein R²    represents C(O)OR⁷ and R⁷ is as hereinbefore defined with    hydroxylamine, or an acid addition salt thereof, for example at room    temperature in the presence of a suitable base (eg potassium    carbonate or triethylamine) and an appropriate organic solvent (eg    THF or EtOH).-   (c) Preparation of a compound of formula I by reaction of a    corresponding compound of formula II,    H—(R)Cgl-Aze-Pab-R²  II    wherein R² is as hereinbefore defined with a compound of formula II,    R¹O(O)C—CH₂-L¹  III    wherein L¹ represents a leaving group, for example halide (eg    bromide) or alkylsulphonate (eg trifluoromethylsulphonate) and R¹ is    as hereinbefore defined, for example between room and elevated    temperature (eg 40° C.) in the presence of a suitable base (eg    potassium carbonate) and an appropriate organic solvent (eg THF, DMF    or acetonitrile).-   (d) Preparation of a compound of formula I wherein R¹ represents H    and R² represents OH or C(O)OR⁷ and R⁷ is as hereinbefore defined by    reaction of a corresponding compound of formula I wherein R¹    represents C₁₋₁₀ alkyl or C₁₋₃ alkylphenyl, and R² represents OH or    C(O)OR⁷, with a suitable base (eg an alkali metal alkoxide or    hydroxide), for example at room temperature in the presence of an    appropriate organic solvent (eg water or MeOH).-   (e) Preparation of a compound of formula I wherein R² represents    OC(O)R⁶ and R⁶ is as hereinbefore defined, by reaction of a    corresponding compound of formula I wherein R² represents OH, with a    compound of formula IV,    R⁶C(O)—O—C(O)R⁶  IV    or a compound of formula V,    R⁶C(O)Hal  V    wherein Hal represents Cl or Br and, in both cases, R⁶ is as    hereinbefore defined, for example at room temperature in the    presence of a suitable base (eg triethylamine, pyridine or DMAP) and    an appropriate organic solvent (eg methylene chloride or THF).-   (f) Preparation of a compound of formula I wherein R¹ represents H    and R² represents OC(O)R⁶, and R⁶ is as hereinbefore defined, by    reaction of a corresponding compound of formula VI    P¹O(O)C—CH₂—(R)Cgl-Aze-Pab-R²  VI    wherein P¹ represents an acid labile ester protecting group (eg tBu    or Bn), and R² represents OC(O)R⁶, wherein R⁶ is as hereinbefore    defined, with a suitable acid (eg TFA), for example at room    temperature in the presence of an appropriate organic solvent (eg    methylene chloride).-   (g) Preparation of a compound of formula I wherein R¹ represents R³,    R³ represents C₁₋₁₀ alkyl or C₁₋₃ alkylphenyl, R² represents OH or    C(O)OR⁷, and R⁷ is as hereinbefore defined, by a transesterification    of a corresponding compound of formula VII,    R^(1a)O(O)C—CH₂—(R)Cgl-Aze-Pab-R²  VII    wherein R^(1a) represents a C₁₋₁₀ alkyl or C₁₋₃ alkylphenyl group    other than that being formed and R² is as hereinbefore defined or an    alternative labile alkyl substituent, under conditions which are    well known to those skilled in the art.

Compounds of formula II may be prepared by deprotection of a compound offormula VIII,Boc-(R)Cgl-Aze-PabR²  VIIIwherein R² is as hereinbefore defined, under conditions which are wellknown to those skilled in the art.

Compounds of formula VI and VII may be prepared analogously to thosemethods described hereinbefore for preparation of compounds of formulaI, in which R¹ represents R³ and R³ represents C₁₋₁₀ alkyl or C₁₋₃alkylphenyl.

Compounds of formula VIII may be prepared by reaction of a compound offormula IX,H-Pab-R²  IXwherein R² is as hereinbefore defined with Boc-Cgl-Aze-OH, for exampleat room temperature in the presence of a suitable coupling system (egEDC), an appropriate base (eg DMAP) and a suitable organic solvent (egdichloromethane or acetonitrle).

Compounds of formula VIII, wherein R² represents OH may be prepared byreaction of a corresponding compound of formula VIII, wherein R²represents C(O)OR⁷ or C(O)OCH(R⁸)OC(O)R⁹, with hydroxylamine, or an acidaddition salt thereof, for example at room temperature in the presenceof a suitable base (eg potassium carbonate or triethylamine) and anappropriate organic solvent (eg THF or EtOH).

Compounds of formula VIII, wherein R² represents C(O)OR⁷ orC(O)OCH(R⁸)OC(O)R⁹, may be prepared by reaction of Boc-(R)Cgl-Aze-Pab-Hwith a compound of formula X,L²C(O)OR^(2a)  Xwherein L² represents a leaving group (eg halogen or phenolate) andR^(2a) represents R⁷ or —CH(R⁸)OC(O)R⁹ and R⁷, R⁸ and R⁹ are ashereinbefore defined, for example at or below room temperature in thepresence of a suitable base (eg NaOH) and an appropriate organic solvent(eg THF).

Compounds of formula VIII, wherein R² represents OC(O)R⁶ mayalternatively be prepared by reaction of a corresponding compound offormula VIII, wherein R² represents OH with a compound of formula IV ashereinbefore defined or a compound of formula V as hereinbefore defined,for example at room temperature in the presence of a suitable base (egtriethylamine, pyridine or DMAP) and an appropriate organic solvent (egmethylene chloride or THF).

Compounds of formula VIII wherein R² represents OC(O)R⁶ mayalternatively be prepared by reaction of Boc-(R)Cgl-Aze-Pab-H with acompound of formula XI,R⁶C(O)—O—O—C(O)R⁶  XIwherein R⁶ is as hereinbefore defined, for example at room temperaturein the presence of an appropriate organic solvent (eg THF).

Compounds of formula VIII wherein R² represents OH may be prepared byreaction of a corresponding compound of formula VIII, wherein R²represents OC(O)R⁶ and R⁶ is as hereinbefore defined with a suitablebase (eg an alkali metal alkoxide), for example at room temperature inthe presence of an appropriate solvent (eg THF).

Compounds of formula IX are well known in the literature or may beprepared using methods analogous to those described hereinbefore. Forexample, compounds of formula IX wherein R² represents C(O)OR⁷ orC(O)OCH(R⁸)OC(O)R⁹ and R⁷, R⁸ and R⁹ are as hereinbefore defined may beprepared by reaction of H-Pab-H, or a protected derivative thereof, witha compound of formula X, as hereinbefore defined, for example at orbelow room temperature in the presence of a suitable base (eg NaOH) andan appropriate organic solvent (eg THF).

Boc-(R)Cgl-Aze-Pab-H may be prepared by reaction of H-Pab-H, or aprotected derivative thereof, with Boc-Cgl-Aze-OH, for example asdescribed hereinbefore for compounds of formula VIII.

Boc-(R)Cgl-Aze-Pab-H may alternatively be prepared by deprotection of acompound of formula XII,Boc-(R)Cgl-Aze-Pab-P²  XIIwherein P² represents a protecting group orthogonal to Boc, underconditions which are well known to those skilled in the art.

Compounds of formula III, IV, V, X, XI and XII are either commerciallyavailable, are well known in the literature, or are available usingknown techniques (eg as described hereinafter).

The compounds of the invention may be isolated from their reactionmixtures using conventional techniques.

It will be appreciated by those skilled in the art that in the processdescribed above the functional groups of intermediate compounds may needto be protected by protecting groups.

Functional groups which it is desirable to protect include hydroxy,amino, amidino and carboxylic acid. Suitable protecting groups forhydroxy include trialkylsilyl and diarylsilyl groups (egt-butyldimethylsilyl, t-butyldiphenylsilyl or trimethylsilyl) andtetrahydropyranyl. Suitable protecting groups for carboxylic acidinclude C₁₋₆ alkyl or benzyl esters. Suitable protecting groups foramino and amidino include t-butyloxycarbonyl or benzoyloxy carbonyl.Amidino nitrogens may be either mono or diprotected.

Protecting groups may be removed in accordance with techniques which arewell known to those skilled in the art, such as those describedhereinafter.

The use of protecting groups is fully described in ‘Protective Groups inOrganic Chemistry’, edited by J W F McOmie, Plenum Press (1973), and‘Protective Groups in Organic Synthesis’, 2nd edition, T W Greene & P GM Wutz, Wiley-Interscience (1991).

Medical and Pharmaceutical Use

The compounds of the invention are useful because they are metabolisedin the body to form compounds which possess pharmacological activity.They are therefore indicated as pharmaceuticals and in particular asprodrugs.

In particular, the compounds of the invention, although they areinactive to thrombin per se, are metabolised in the body to form potentinhibitors of thrombin, for example as demonstrated in the testdescribed below.

By “the compounds of the invention are inactive to thrombin per se” wemean that they exhibit an IC₅₀TT value, as determined in Test A below,of greater than 1 μM.

The compounds of the invention are thus expected to be useful in thoseconditions where inhibition of thrombin is required.

The compounds of the invention are thus indicated both in thetherapeutic and/or prophylactic treatment of thrombosis andhypercoagulability in blood and tissues of animals including man.

It is known that hypercoagulability may lead to thrombo-embolicdiseases. Thrombo-embolic diseases which may be mentioned includeactivated protein C resistance, such as the factor V-mutation (factor VLeiden), and inherited or acquired deficiencies in antithrombin m,protein C, protein S, heparin cofactor II. Other conditions known to beassociated with hypercoagulability and thrombo-embolic disease includecirculating antiphospholipid antibodies (Lupus anticoagulant),homocysteinemi, heparin induced thrombocytopenia and defects infibrinolysis. The compounds of the invention are thus indicated both inthe therapeutic and/or prophylactic treatment of these conditions.

The compounds of the invention are further indicated in the treatment ofconditions where there is an undesirable excess of thrombin withoutsigns of hypercoagulability, for example in neurodegenerative diseasessuch as Alzheimer's disease.

Particular disease states which may be mentioned include the therapeuticand/or prophylactic treatment of venous thrombosis and pulmonaryembolism, arterial thrombosis (eg in myocardial infarction, unstableangina, thrombosis-based stroke and peripheral arterial thrombosis) andsystemic embolism usually from the atrium during arterial fibrillationor from the left ventricle after transmural myocardial infarction.

Moreover, the compounds of the invention are expected to have utility inprophylaxis of re-occlusion (ie thrombosis) after thrombolysis,percutaneous trans-luminal angioplasty (PTA) and coronary bypassoperations; the prevention of re-thrombosis after microsurgery andvascular surgery in general.

Further indications include the therapeutic and/or prophylactictreatment of disseminated intravascular coagulation caused by bacteria,multiple trauma, intoxication or any other mechanism; anticoagulanttreatment when blood is in contact with foreign surfaces in the bodysuch as vascular grafts, vascular stents, vascular catheters, mechanicaland biological prosthetic valves or any other medical device; andanticoagulant treatment when blood is in contact with medical devicesoutside the body such as during cardiovascular surgery using aheart-lung machine or in haemodialysis.

In addition to its effects on the coagulation process, thrombin is knownto activate a large number of cells (such as neutrophils, fibroblasts,endothelial cells and smooth muscle cells). Therefore, the compounds ofthe invention may also be useful for the therapeutic and/or prophylactictreatment of idiopathic and adult respiratory distress syndrome,pulmonary fibrosis following treatment with radiation or chemotherapy,septic shock, septicemia, inflammatory responses, which include, but arenot limited to, edema, acute or chronic atherosclerosis such as coronaryarterial disease, cerebral arterial disease, peripheral arterialdisease, reperfusion damage, and restenosis after percutaneoustrans-luminal angioplasty (PTA).

Compounds of the invention that inhibit trypsin and/or thrombin may alsobe useful in the treatment of pancreatitis.

According to a further aspect of the present invention, there isprovided a method of treatment of a condition where inhibition ofthrombin is required which method comprises administration of atherapeutically effective amount of a compound of formula I as definedabove, or a pharmaceutically acceptable salt thereof, to a personsuffering from, or susceptible to such a condition.

The compounds of the invention will normally be administered orally,buccally, rectally, dermally, nasally, tracheally, bronchially, by anyother parenteral route or via inhalation, in the form of pharmaceuticalpreparations comprising the prodrug either as a free base, or apharmaceutical acceptable non-toxic organic or inorganic acid additionsalt, in a pharmaceutically acceptable dosage form. Depending upon thedisorder and patient to be treated and the route of administration, thecompositions may be administered at varying doses.

The compounds of the invention may also be combined and/orco-administered with any antithrombotic agent with a different mechanismof action, such as the antiplatelet agents acetylsalicylic acid,ticlopidine, clopidogrel, thromboxane receptor and/or synthetaseinhibitors, fibrinogen receptor antagonists, prostacyclin mimetics andphosphodiesterase inhibitors and ADP-receptor (P₂T) antagonists.

The compounds of the invention may further be combined and/orco-administered with thrombolytics such as tissue plasminogen activator(natural or recombinant), streptokinase, urokinase, prourokinase,anisolated streptokinase plasminogen activator complex (ASPAC), animalsalivary gland plasminogen activators, and the like, in the treatment ofthrombotic diseases, in particular myocardial infarction.

According to a further aspect of the invention there is thus provided apharmaceutical formulation including a compound of formula I ashereinbefore defined, or a pharmaceutically acceptable salt thereof, inadmixture with a pharmaceutically acceptable adjuvant, diluent orcarrier.

Suitable daily doses of the compounds of the invention in therapeuticaltreatment of humans are about 0.001-100 mg/kg body weight at peroraladministration and 0.001-50 mg/kg body weight at parenteraladministration.

The compounds of the invention have the advantage that they may haveimproved pharmacokinetic properties, such as those identifiedhereinbefore, both after oral and parenteral administration, whencompared with compounds of formula:R^(a)(O)C—CH₂—(R)Cgl-Aze-Pab-Hwherein R^(a) is as hereinbefore defined, and in particular the compoundwherein R^(a) represents H.

The compounds of the invention are inactive to thrombin, trypsin andother serine proteases. The compounds thus remain inactive in thegastrointestinal tract and the potential complications experienced byorally administered anticoagulants which are active per se, such asbleeding and indigestion resulting from inhibition of trypsin, may thusbe avoided.

Furthermore, local bleeding associated with and after parenteraladministration of an active thrombin inhibitor may be avoided by usingthe compounds of the invention.

The compounds of the invention may also have the advantage that they maybe more efficacious than, be less toxic than, be longer acting than,have a broader range of activity than, produce fewer side effects than,be more easily absorbed than, or that they may have other usefulpharmacological properties over, compounds known in the prior art.

Biological Tests

Test A

Determination of Thrombin Clotting Time (TT)

Human thrombin (T 6769, Sigma Chem Co, final concentration of 1.4 NIHunits/mL) in buffer solution, pH 7.4, 100 μL, and inhibitor solution,100 μL, were incubated for one min. Pooled normal citrated human plasma,100 μL, was then added and the clotting time measured in an automaticdevice (KC 10, Amelung).

The clotting time in seconds was plotted against the inhibitorconcentration, and the IC₅₀TT was determined by interpolation.

IC₅₀TT is the concentration of inhibitor that doubles the thrombinclotting time for human plasma.

Test B

Determination of Thrombin Time in Plasma Ex Vivo

The inhibition of thrombin after oral or parenteral administration ofthe compounds of the invention were examined in conscious rats which,one or two days prior to the experiment, were equipped with a catheterfor blood sampling from the carotid artery. On the experimental day, thecompound, dissolved in ethanol:Solutol™:water (5:5:90), was administeredand blood samples were withdrawn at fixed times into plastic tubescontaining 1 part sodium citrate solution (0.13 mol per L.) and 9 partsof blood. The tubes were centrifuged to obtain platelet poor plasma. Theplasma was used for determination of thrombin time as described below.

The citrated rat plasma, 100 μL, was diluted with a saline solution,0.9%, 100 μL, and plasma coagulation was started by the addition ofhuman thrombin (T 6769, Sigma Chem Co, USA) in a buffer solution, pH7.4, 100 μL. The clotting time was measured in an automatic device (KC10, Amelumg, Germany).

The concentrations of the active thrombin inhibitorHO(O)C—CH₂(R)Cgl-Aze-Pab-H (see International Patent Application WO94/29336) in the rat plasma were estimated by the use of standard curvesrelating the thrombin time in the pooled citrated rat plasma to knownconcentrations of the aforementioned active thrombin inhibitor dissolvedin saline.

Based on the estimated plasma concentrations of the active thrombininhibitor HO(O)C—CH₂(R)Cgl-Aze-Pab-H (which assumes that thrombin timeprolongation is caused by the aforementioned compound) in the rat, thearea under the curve after oral and/or parenteral administration of theprodrug was calculated (AUCpd) using the trapezoidal rule andextrapolation of data to infinity.

The bioavailability of the active thrombin inhibitorHO(O)C—CH₂(R)Cgl-Aze-Pab-H after oral or parenteral administration ofthe prodrug was calculated as below:[(AUCpd/dose)/(AUCactive,iv/dose]×100where AUCactive,iv represents the AUC obtained after intravenousadministration of HO(O)C—CH₂(R)Cgl-Aze-Pab-H to conscious rats asdescribed above.Test CDetermination of Thrombin Time in Urine Ex Vivo

The amount of the active thrombin inhibitor HO(O)C—CH₂(R)Cgl-Aze-Pab-Hthat was excreted in urine after oral or parenteral administration ofthe compounds of the invention, dissolved in ethanol:Solutol™:water(5:5:90), was estimated by determination of the thrombin time in urineex vivo (assuming that thrombin time prolongation is caused by theaforementioned compound).

Conscious rats were placed in metabolism cages, allowing separatecollection of urine and faeces, for 24 hours following oraladministration of compounds of the invention. The thrombin time wasdetermined on the collected urine as described below.

Pooled normal citrated human plasma (100 μL) was incubated with theconcentrated rat urine, or saline dilutions thereof, for one minute.Plasma coagulation was then initiated by the administration of humanthrombin (T 6769, Sigma Chem Company) in buffer solution (pH 7.4; 100μL). The clotting time was measured in an automatic device (KC 10;Amelung).

The concentrations of the active thrombin inhibitorHO(O)C—CH₂(R)Cgl-Aze-Pab-H in the rat urine were estimated by the use ofstandard curves relating the thrombin time in the pooled normal citratedhuman plasma to known concentrations of the aforementioned activethrombin inhibitor dissolved in concentrated rat urine (or salinedilutions thereof). By multiplying the total rat urine production overthe 24 hour period with the estimated mean concentration of theaforementioned active inhibitor in the urine, the amount of the activeinhibitor excreted in the urine (AMOUNTpd) could be calculated.

The bioavailability of the active thrombin inhibitorHO(O)C—CH₂(R)Cgl-Aze-Pab-H after oral or parenteral administration ofthe prodrug was calculated as below:[(AMOUNTpd/dose)/(AMOUNTactive,iv/dose]×100where AMOUNTactive,iv represents the amount excreted in the urine afterintravenous administration of HO(O)C—CH₂(R)Cgl-Aze-Pab-H to consciousrats as described above.Test DDetermination of HO(O)C—CH₂—(R)Cgl-Aze-Pab-H in Urine by LC-MS

The amount of the active thrombin inhibitor HO(O)C—CH₂—(R)Cgl-Aze-Pab-Hthat was excreted in urine after oral or parenteral administration ofthe compounds of the invention, dissolved in ethanol:Solutol™:water(5:5:90), was measured by LC-MS analysis as described below.

The animal studies were performed as described in Method C above. Urinesamples were collected and frozen at −20° C. before they were analysed.

Urine samples were analysed for their content ofHO(O)C—CH₂—(R)Cgl-Aze-Pab-H according to the following method:

Thawed urine samples were mixed and, if required, spinned in acentrifuge. Solid phase extraction tubes (Analytichem Bond Elut. No.1210-2059) were activated with 1.0 mL of methanol and conditioned with1.0 mL of acetonitrile:water (50:50), followed by 1.0 mL of 0.1% formicacid. 50 μL of the working internal standard (20 μmol/L) was added toeach extraction tube. For urine standards, 50 μL of standard solutionwas added. 200 μL of a sample or, for urine standards, blank urine wasadded to each tube and thereafter pulled through via gravity or a gentlevacuum. Residual urine was washed out with 1.0 mL of ammonium acetate (2mmol/L), before elution with 1.0 mL of acetonitrile:ammonium acetate (2mmol/L) (35:65). The collected eluate was transferred to autosamplervials. 30 μL of the extract was injected onto the LC column (HypersilBDS-C18; 3 μm; 75 mm×4.0 mm i.d.; Hewlett-Packard No. 79926 03-354),eluted with ammonium acetate buffer (1.3 mmol/L) with 40% acetonitrileand 0.1% formic acid at 0.75 mL/min. The effluent was split so that 30μL/min entered the electrospray ion source of a P-E Sciex API-3 massspectrometer. HO(O)C—CH₂—(R)Cgl-Aze-Pab-H andHO(O)C—CH₂—(R)Cgl-Pro-Pab-H (internal standard) both have retentiontimes near 1.5 minutes. Their molecular ions ((M+H)⁺) were monitored atm/z 430.2 and 444.2 respectively, at unit mass resolution. Urinestandards at two levels, one being at the limit of quantification, wereused for calibration based on peak area ratios ofHO(O)C—CH₂—(R)Cgl-Aze-Pab-H over the internal standard. Linearity of themethod was checked over the range 0.050-20 μmol/L. The coefficient ofvariation was 1-2% at 1-20 μmol/L and 7% at 0.50 μmol/L. The limit ofquantification was 0.050 μmol/L.

By multiplying the total urine production over the 24 hour period by themeasured concentration of HO(O)C—CH₂—(R)Cgl-Aze-Pab-H in the urine, theamount of the active inhibitor excreted in urine (AMOUNTpd) could becalculated. The bioavailability of the active thrombin inhibitor wasthen calculated as described in Method C above.

The invention is illustrated by way of the following examples.

EXAMPLES General Experimental Procedures

Mass spectra were recorded on a Finnigan MAT TSQ 700 triple quadrupolemass spectrometer equipped with an electrospray interface.

The ¹H NMR and ¹³C NMR measurements were performed on BRUKER ACP 300 andVarian UNITY plus 400 and 500 spectrometers, operating at ¹H frequenciesof 300.13, 399.96 and 499.82 MHz respectively, and at ¹³C frequencies of75.46, 100.58 and 125.69 MHz respectively. Chemical shifts are reportedin δ units.

Preparation of Starting Materials

Boc-(R)Cgl-Aze-Pab-H, Boc-(R)Cgl-Aze-Pab×HCl, H—(R)Aze-Pab-Z,H—(R)Aze-Pab-Z×HCl, Bn-OOCCH₂—(R)Cgl-Aze-Pab-Z, Boc-(R)Cgl-Aze-Pab-Z,Boc-(R)Cgl-Aze-OH and Pab-Z×HCl were prepared according to the methodsdescribed in International Patent Application WO 94/29336.

EXAMPLE 1 EtOOCCH₂—(R)Cgl-Aze-Pab-COOCH₂CH═CH₂

(i) Boc-(R)Cgl-Aze-Pab-COOCH₂CH═CH₂

To a solution of Boc-(R)Cgl-Aze-Pab-H (6.1 g; 13 mmol) in THF (125 mL)and 2M NaOH (70 mL; 140 mmol) at 0° C. was added dropwise allylchloroformate (1.7 g; 14 mmol). After stirring at 0° C. for 1 h, thereaction was mixture concentrated, water was added (100 mL) and theresulting aqueous phase was extracted with methylene chloride (3×100mL). The combined organic phases were concentrated to give 6.4 g of acrude product which was purified by flash chromatography usingEtOAc:THF:Et₃N (68:29:3) as eluent. Concentration gave 5.8 g (81%) ofthe subtitle compound as a white solid.

¹H NMR (500 MHz, CDCl₃): δ 8.19 (bt, 1H), 7.78 (d, 2H), 7.26 (d, 2H),6.02-5.92 (m, 1H), 5.32 (d, J=17 Hz, 1H), 5.18 (d, J=10 Hz, 1H), 5.06(d, J=7 Hz, 1H), 4.82 (bs, 1H), 4.61 (d, J=6 Hz, 2H), 4.58-4.48 (m, 1H),4.38-4.27 (m, 2H), 4.14-4.03 (m, 1H), 3.77-3.68 (m, 1H), 2.60-0.90 (m,24H). ¹³C NMR (125 MHz, CDCl₃) carbonyl and amidine signals: δ 172.70,170.74, 168.02, 164.54, 155.98.

(ii) H—(R)Cgl-Aze-Pab-COOCH₂CH═CH₂×2TFA

To a solution of Boc-(R)Cgl-Aze-Pab-COOCH₂CH═CH₂(2.03 g; 3.65 mmol; fromstep (i) above) in methylene chloride (15 mL) at 0° C. was added TFA (15mL). The reaction mixture was stirred at ambient temperature for 3 hfollowed by concentration to give the 2.8 g of the subtitle compound asa white solid.

¹H NMR (500 MHz, MeOH (d4)): δ 7.80 (d, 2H), 7.57 (d, 2H), 6.02 (m, 1H),5.45 (d, J=17 Hz, 1H), 5.33 (d, J=10 Hz, 1H), 5.91-4.80 (m, 3H), 4.56(s, 2H), 4.38 (bq, J=8 Hz, 1H), 3.71 (d, J=7 Hz, 1H), 2.76-2.60 (m, 1H),2.35-2.20 (m, 1H), 1.9-1.0 (m, 11H).

(iii) EtOOCCH₂—(R)Cgl-Aze-Pab-COOCH₂CH═CH₂

A mixture of H—(R)Cgl-Aze-Pab-COOCH₂CH═CH₂×2TFA (649 mg; 0.95 mmol; fromstep (ii) above), K₂CO₃ (656 mg, 4.8 mmol), water (0.1 mL), and THF (10mL) was stirred at 40° C. for 2 h followed by addition of ethylbromoacetate (190 mg; 1.14 mmol) in TH (1 mL). After stirring at 40° C.for 4 h and at ambient temperature for 14 h the reaction mixture wasfiltered, concentrated, and purified by flash chromatography usingEtOAc:THF:Et₃N (68:29:3) as eluent to give 244 mg (47%) of the titlecompound as a white solid.

¹NMR (400 MHz, CDCl₃): δ 8.46 (bt, 1H), 7.81 (d, 2H), 7.35 (d, 2H),6.08-5.94 (m, 1H), 5.35 (d, J=18 Hz, 1H), 5.23 (d, J=11 Hz, 1H), 4.93(dd, J=6 and 9 Hz, 1H), 4.66 (d, 2H), 4.62-4.38 (AB part of anABX-spectrum), 4.16-4.04) (m, 4H), 3.20 (d, 2H), 2.86 (d, 1H), 2.64-2.45(m, 2H), 2.0-1.0 (m 17H). ¹³C NMR (100 MHz, CDCl₃) carbonyl and amidinesignals: δ 175.33, 172.24, 170.72, 168.19, 164.35.

EXAMPLE 2 nPrOOCCH₂—(R)Cgl-Aze-Pab-COOCH₂CH═CH₂

The title compound was prepared according to the procedure described inExample 1(iii) from H—(R)Cgl-Aze-Pab-COOCH₂CH═CH₂×2TFA (503 mg; 0.74mmol; see Example 1(ii) above) and n-propyl bromoacetate (160 mg, 0.88mmol) to give 277 mg (68%) as a white solid.

¹H-NMR (400 MHz, CDCl₃): δ 8.48 (bt, 1H), 7.83 (d, 2H), 7.35 (d, 2H),6.76 (broad, 1H), 6.02 (m, 1H), 5.37 (dd, 1H), 5.24 (dd, 1H), 4.94 (t,1H), 4.67 (dd, 2H), 4.49 (AB part of an ABX-spectrum, 2H), 4.12 (m, 2H),3.98 (t, 2H), 3.24 (AB-system, 2H), 2.87 (d, 1H), 2.52 (m, 2H), 1.99(bd, 2H), 1.80-1.50 (m, 7H), 1.61 (q, 2H), 1.30-1.10 (m, 2H), 1.00 (qd,2H), 0.90 (t, 3H). ¹³C-NMR (100 MHz, CDCl₃) amidine and carbonylsignals: δ 175.4, 172.3, 170.7, 167.9, 164.5

EXAMPLE 3 tBuOOCCH₂—(R)Cgl-Aze-Pab-COOCH₂CH═CH₂

The title compound was prepared according to the procedure described inExample 1(iii) from H—(R)Cgl-Aze-Pab-COOCH₂CH═CH₂×2TFA (285 mg; 0.42mmol; see Example 1(ii) above) and t-butyl bromoacetate (96 mg; 0.50mmol) to give 93 mg (39%) as a white solid.

¹H NMR (500 MHz, CDCl₃): δ 8.50 (bt, 1H), 7.81 (d, 2H), 7.36 (d, 2H),6.07-5.97 (m, 1H), 5.36 (d, J=16 Hz, 1H), 5.22 (d, J=10 Hz, 1H), 4.93(dd, J=9 and 6 Hz, 1H), 4.76 (d, J=6 Hz, 2H), 4.57-4.46 (m, 2H),4.18-4.04 (m, 2H), 3.19-3.08 (AB-spectrum, J_(AB)=20 Hz, 2H), 2.86 (d,J=8 Hz, 1H), 2.72-2.53 (m, 2H), 2.0-0.9 (m, 23H). ¹³C NMR (100 MHz,CDCl₃) amidine and carbonyl signals: δ 175.28, 171.53, 170.76, 167.81,164.1.

EXAMPLE 4 EtOOCCH₂—(R)Cgl-Aze-Pab-COOEt

(i) Boc-(R)Cgl-Aze-Pab-COOEt

The sub-title compound was prepared according the procedure described inExample 1(i) from Boc-(R)Cgl-Aze-Pab-H (600 mg; 1.3 mmol) and ethylchloroformate (150 mg; 1.4 mmol) yielding 240 mg (34%) as a white solid.

¹H-NMR (300 MHz, CDCl₃): δ 9.37 (bs, 1H), 8.16 (bs, 1H), 7.72 (d, 2H),7.18 (d, 2H), 5.17 (d, 1H), 4.73 (t, 1H), 4.47 (dd, 1H), 4.27 (m, 2H),4.06 (q, 2H), 3.66 (t, 1H), 2.48 (m, 1H), 2.37 (m, 1H), 1.4-1.8 (m, 7H),1.22 (s, 9H), 1.3-0.8 (m, 7H). ¹³C-NMR (75 MHz, CDCl₃) carbonyl andamidine signals: δ 172.6, 170.7, 167.9, 164.8, 156.0

(ii) H—(R)Cgl-Aze-Pab-COOEt×2HCl

To a solution of Boc-(R)Cgl-Aze-Pab-COOEt (240 mg; 0.44 mmol; from step(i) above) in EtOAc (20 mL) was added hydrogen chloride at 0° C. over 5minutes. The reaction mixture was stirred at 0° C. for 1 h followed byconcentration to give 225 mg (100%) as a white solid.

¹H-NMR (300 MHz, D₂O): δ 7.85 (d, 2H), 7.61 (d, 2H), 4.98 (dd, 1H), 4.60(s, 1H), 4.44 (p, 5H), 3.90 (d, 1H), 2.73 (m, 1H), 2.37 (m, 1H),2.0-1.65 (m, 9H), 1.39 (t, 3H), 1.4-1.1 (m, 7H), 0.98 (m, 1H). ³C-NMR(75 MHz, D₂O) amidine and carbonyl signals: δ 172.7, 169.4, 166.8,154.3.

(iii) EtOOCCH₂—(R)Cgl-Aze-Pab-COOEt

The title compound was prepared according to the procedure described inExample 1(iii) from H—(R)Cgl-Aze-Pab-COOEt×2HCl (160 mg; 0.31 mmol; fromstep (ii) above) and ethyl bromoacetate (52.5 mg; 0.31 mmol). Yield: 100mg (61%) as a light yellow powder.

¹H-NMR (300 MHz, CDCl₃): δ 8.48 (bt, 1H), 7.81 (d, 2H), 7.38 (d, 2H),4.51 (AB part of an ABX-spectrum, 2H), 4.21 (q, 2H), 4.15-4.05 (m, 4H),3.21 (AB-spectrum, 2H), 2.86 (d, 1H), 2.68 (m, 1H), 2.53 (m, 1H), 1.96(bd, 2H), 1.90-1.70 (m, 12H), 1.35 (t, 3H), 1.22 (t, 6H), 1.30-0.95 (m,2H). ¹³C-NMR (75 MHz, CDCl₃) carbonyl and amidine signals: δ 175.5,172.2, 170.7, 167.6, 164.9

EXAMPLE 5 EtOOCCH—(R)Cgl-Aze-Pab-COO-nPr

(i) Boc-(R)Cgl-Aze-Pab-COO-nPr

The sub-title compound was prepared according to the procedure describedin Example 1(i) above using Boc-(R)Cgl-Aze-Pab-H (6.0 g; 13 mmol;) andn-propyl chloroformate (1.57 mL; 14 mmol). Yield 5.4 g (76%).

¹H-NMR (400 MHz, CDCl₃): δ 8.25 (bt, 1H), 7.82 (d, 2H), 7.31 (d, 2H),5.09 (bd, 1H), 4.87 (dd, 1H), 4.58 (dd, 1H), 4.39 (dd, 2H), 4.14 (q,1H), 4.10 (t, 2H), 3.79 (t, 1H), 2.54 (dm, 2H), 2.21 (s, 1H), 1.87-1.55(m, 8H), 1.33 (s, 9H), 1.45-1.0 (m, 4H), 0.99 (t, 3H). ¹³C-NMR (100 MHz,CDCl₃) amidine and carbonyl signals: δ 172.7, 170.6, 167.8, 165.0,155.9.

(ii) H—(R)Cgl-Aze-Pab-COO-nPr×2TFA

The sub-title compound was prepared according to the procedure describedin Example 1(ii) using 2.1 g (3.7 mmol) of Boc-(R)Cgl-Aze-Pab-COO-nPr(from step (i) above). Yield 3.7 g.

¹H-NMR (400 MHz, MeOH-d₄): δ 7.77 (d, 2H), 7.60 (d, 1H), 4.86 (dd, 1H),4.56 (AB part of an ABX-spectrum, 2H), 4.33 (m, 4H), 3.72 (d, 1H), 3.30(m, 1H), 2.68 (m, 1H), 2.28 (m, 1H), 1.9-1.7 (m, 9H), 1.4-1.1 (m, 6H),1.02 (t, 3H). ¹³C-NMR (100 MHz, MeOH-d₄) carbonyl and amidine signals: δ172.7, 169.3, 168.0, 161.4.

(iii) EtOOCCH₂—(R)Cgl-Aze-Pab-COO-nPr

The title compound was prepared according to the procedure described inExample 1(iii) from H—(R)Cgl-Aze-Pab-COO-nPr×2TFA (472 mg; 0.69 mmol;from step (ii) above) and ethyl bromoacetate (138 mg; 0.83 mmol) to give0.22 mg (58%) as a white solid.

¹H-NMR (400 MHz, CDCl₃): δ 8.46 (bt, 1H), 7.82 (d, 2H), 7.32 (d, 2H),4.92 (dd, 1H), 4.49 (AB part of an ABX-spectrum, 2H), 4.10 (m, 6H), 3.23(AB-spectrum, 2H), 2.80 (dm, 2H), 1.98 (bd, 2H), 1.74 (q, 2H), 1.63 (dd,2H), 1.52 (m, 1), 1.21 (t, 3H), 1.20-1.10 (m, 2H), 0.98 (t, 3H). ¹³C-NMR(100 MHz, CDCl₃) carbonyl and amidine signals: δ 175.3, 172.2, 170.7,167.6, 164.3.

EXAMPLE 6 MeOOCCH₂—(R)Cgl-Aze-Pab-COO-nPr

The title compound was prepared according to the procedure described inis Example 1(iii) above from H—(R)Cgl-Aze-Pab-COO-nPr×2TFA (365 mg; 0.53mmol; see Example 5(ii) above) and methyl bromoacetate (98 mg; 0.64mmol) to give 114 mg (41%) as a white solid.

¹H-NMR (500 MHz, CDCl₃): δ 8.44 (bt, 1H), 7.82 (d, 2H), 7.32 (d, 2H),7.04 (broad, 1H), 4.92 (dd, 1H), 4.49 (AB part of an ABX spectrum), 4.12(m, 2H), 4.10 (t, 2H), 3.63 (s, 3H), 3.24 (s, 2H), 2.87 (d, 1H), 2.65(m, 1H), 2.52 (m, 1H), 2.01 (broad, 1H), 1.96 (bd, 2H), 1.75 (q, 4H),1.63 (bdd, 1H), 1.53 (m, 1H), 1.3-1.1 (m, 5H), 0.99 (t, 3H). ¹³C-NMR(100 MHz, CDCl₃) carbonyl and amidine signals: δ 175.3, 172.5, 170.7,167.7, 165.0.

EXAMPLE 7 EtOOCCH₂—(R)Cgl-Aze-Pab-COOCH₂CH₂OMe

(i) Boc-(R)Cgl-Aze-Pab-COOCH₂CH₂OMe

The sub-title compound was prepared according to the procedure describedin Example 1(i) above using Boc-(R)Cgl-Aze-Pab-H (6.0 g; 13 mmol) and2-methoxyethyl chloroformate (1.94 g; 14 mmol). Yield 3.9 g (52%).

¹H-NMR (400 MHz, CDCl₃): δ 8.24 (bt, 1H), 7.83 (d, 2H), 7.31 (d, 2H),5.08 (bd, 1H), 4.87 (dd, 1H), 4.58 (dd, 1H), 4.39 (dd, 2H), 4.30 (t,2H), 4.15 (m, 1H), 3.79 (bt, 1H), 3.68 (t, 2H), 3.40 (s, 3H), 2.65-2.45(m, 2H), 2.20 (broad, 1H), 1.9-1.55 (m, 6H), 1.34 (s, 9H), 1.3-0.95 (m,6H). ¹³C-NMR (100 MHz, CDCl₃) carbonyl and amidine signals: δ 172.7,170.7, 167.8, 164.6, 155.9.

(ii) H—(R)Cgl-Aze-Pab-COOCH₂CH₂OMe×2TFA

The sub-title compound was prepared according to the procedure describedin Example 1(ii) above using 1.71 g of Boc-(R)Cgl-Aze-Pab-COOCH₂CH₂OMe(from step (i) above). Yield 1.89 g (88%).

¹H-NMR (400 MHz, MeOH-d4): δ 7.77 (d, 2H), 7.59 (d, 2H), 4.85 (dd, 1H),4.56 (d, 2H), 4.49 (m, 2H), 4.37 (m, 1H), 4.28 (m, 1H), 3.70 (m, 3H),3.37 (s, 3H), 2.68 (m, 1H), 2.28 (m, 1H), 1.9-1.7 (m, 7H), 1.4-1.1 (m,6H). ¹³C-NMR (100 MHz, MeOH-d4) carbonyl and amidine signals: δ 172.7,169.3, 168.0, 154.6.

(iii) EtOOCCH₂—(R)Cgl-Aze-Pab-COOCH₂CH₂OMe

The title compound was prepared according to the procedure described inExample 1(iii) above from H—(R)Cgl-Aze-Pab-COOCH₂CH₂OMe×2TFA (487 mg;0.69 mmol; from step (ii) above) and ethyl bromoacetate (138 mg; 0.83mmol) to give a crude product which was purified by flash chromatographyusing THF:methylene chloride (3:1) as eluent. The yield was 0.13 mg(34%) as a white solid.

¹H-NMR (400 MHz, CDCl₃): δ 8.46 (bt, 1H), 7.83 (d, 2H), 7.32 (d, 2H),7.21 (broad, 1H), 4.92 (dd, 1H), 4.49 (AB part of an ABX spectrum, 2H),4.30 (t, 2H), 4.12 (q, 2H), 4.07 (q, 2H), 3.68 (t, 1H), 3.40 (s, 3H),3.24 (s, 2H), 2.62 (m, 1H), 2.52 (m, 1H), 2.07 (broad, 1H), 1.97 (bd,1H), 1.8-1.5 (m, 5H), 1.3-1.1 (m, 6H), 1.05-4.95 (m, 2H). ¹³C-NMR (100MHz, CDCl₃) carbonyl and amidine signals: δ 175.3, 172.2, 170.7, 167.8,164.6.

EXAMPLE 8 MeOOCCH₂—(R)Cgl-Aze-Pab-COOCH₂CH₂OMe

The title compound was prepared according to the method described inExample 1(iii) above from H—(R)Cgl-Aze-Pab-COOCH₂CH₂OMe×2TFA (490 mg;0.7 mmol; see Example 7(ii) above) and methyl bromoacetate (128 mg; 0.84mmol) to give a crude product which was purified by flash chromatographyusing THF:methylene chloride (3:1) as eluent. The yield was 155 mg (41%)as a white solid.

¹H-NMR (400 MHz, CDCl₃): δ 8.44 (t, 1H), 7.83 (d, 2H), 7.31 (d, 2H),4.92 (dd, 1H), 4.49 (AB part of an ABX spectrum, 2H), 4.30 (t, 2H), 4.13(m, 2H), 3.68 (t, 2H), 3.63 (s, 3H), 3.39 (s, 3H), 3.25 (s, 2H), 2.87(d, 1H), 2.62 (m, 1H), 2.52 (m, 1H), 1.96 (bd, 1H), 1.8-1.5 (m, 6H),1.3-1.1 (m, 5H), 1.00 (q, 2H). ¹³C-NMR (100 MHz, CDCl₃) carbonyl andamidine signals: δ 175.2, 172.6, 170.7, 167.8, 164.5.

EXAMPLE 9 EtOOCCH₂—(R)Cgl-Aze-Pab-COO-nBu

(i) Boc-(R)Cgl-Aze-Pab-COO-nBu

The sub-title compound was prepared according to the procedure describedin Example 1(i) from Boc-(R)Cgl-Aze-Pab-H (1.01 g; 2.1 mmol) and n-butylchloroformate (0.32 g; 2.4 mmol). After stirring at ambient temperaturefor 1.5 h the reaction mixture was concentrated and extracted with threeportions of methylene chloride. The combined organic phase was thenwashed with water, dried over Na₂SO₄, and concentrated to give 1.0 g(83%) of the sub-title compound as a white solid.

¹H-NMR (300 MHz, CDCl₃): δ 9.81-9.31 (bs, 1H), 8.36-8.20 (m, 1H), 7.35(d, 2H), 7.84 (d, 2H), 6.78-6.43 (bs, 1H), 5.05-4.82 (m, 2H), 4.69-4.15(m, 3H), 4.15-4.08 (m, 3H), 3.86-3.70 (m, 1H), 2.68-2.42 (m, 2H),1.92-0.88 (m, 25H). ¹³C-NMR (125 MHz, CDCl₃) amidine and carbonylsignals: δ 172.5, 170.7, 167.9, 164.9, 156.0. FAB-MS: (m+1)=572 (m/z)

(ii) H—(R)Cgl-Aze-Pab-COO-nBu×2HCl

The sub-title compound was prepared according to the procedure describedin Example 4(ii) from Boc-(R)Cgl-Aze-Pab-COO-nBu (2.5 g; 4.4 mmol; fromstep (i) above) to give 2.4 g (100%) as a white solid.

¹H-NMR (300 MHz, MeOH-d4): δ 7.78-7.60 (m, 2H), 4.66-4.49 (m, 2H), 0.98(t, 2H), 4.49-4.35 (m, 3H), 4.35-4.22 (m, 1H), 3.75 (d, 1H), 1.92-1.67(m, 8H), 1.56-1.07 (m, 8H). The signal of one of the protons ispartially obscured by the CD₃OH-signal ¹³C-NMR (100 MHz, MeOH-d4)amidine and carbonyl signals: δ 172.7, 169.3, 167.9, 154.7 MS (m+1)=472(m/z)

(iii) EtOOCCH₂—(R)Cgl-Aze-Pab-COO-nBu

The title compound was prepared analogously to the procedure describedin Example 1(iii) from H—(R)Cgl-Aze-Pab-COO-nBu×2HCl (400 mg; 0.74 mmol)and ethyl bromoacetate (147 mg; 0.88 mmol). The product was purified byflash chromatography using methylene chloride and EtOH gradient0.1%=>12.8% as eluent to give 290 mg (70%) as a white solid.

¹H-NMR (300 MHz, CDCl₃): δ 9.70-9.36 (bs, 1H), 8.47 (t, 1H), 7.81 (d,2H), 7.32 (d, 2H), 7.07-6.73 (bs, 1H), 4.97-4.87 (dd, 1H), 4.62-4.35 (m,2H), 4.20-3.98 (m, 6H), 3.27-3.12 (m, 2H), 2.84 (s, 1H), 2.70-2.40 (m,2H), 2.03-0.85 (m, 22H) ¹³C-NMR (75 MHz, CDCl₃) amidine and carbonylsignals: δ 175.3, 172.3, 170.8, 167.9, 165.0 FAB-MS: (m+1)=558 (m/z)

EXAMPLE 10 PrlC(O)CH₂CH₂CH₂OOCCH₂—(R)Cgl-Aze-Pab-Z

(i) PrlC(O)CH₂CH₂CH₂OH

A mixture of γ-butyrolactone (4.0 g; 46.5 mmol) and pyrrolidine (6.6 g;92.8 mmol) was stirred at room temperature for 2.5 h. The product wasconcentrated in vacuum to give 14.5 g (100%) of the product as a yellowoil.

¹H-NMR (300 MHz, MeOH-d4): δ 3.58 (t, 2H), 3.50 (t, 2H), 3.40 (t, 2H),2.42 (t, 2H), 2.06-1.75 (m, 6H)

(ii) PrlC(O)CH₂CH₂OOCCH₂Br

To a mixture of PrlC(O)CH₂CH₂CH₂OH (7.2 g; 45.8 mmol; from step (i)above) and DMAP (5.6 g; 45.8 mmol) in methylene chloride at 0° C. wasadded dropwise bromoacetyl bromide (4.0 mL; 45.8 mmol). After stirringat room temperature for 1.5 h another portion of bromoacetyl bromide(1.0 mL, 11.4 mmol) and DMAP (1.4 g, 11.4 mmol) was added and reactionwas refluxed for 1.5 h. Water was added and the methylene chloride wasextracted 3 times. The organic phase was dried with Na₂SO₄ andconcentrated to give 10.3 g (81%) of the product as a yellow oil.

¹H-NMR (400 MHz, CDCl₃): δ 4.15 (t, 2H), 3.75 (s, 2H), 3.40-3.31 (m,4H), 2.30 (t, 2H), 1.98-1.83 (m, 4H), 1.81-1.73 (m, 2H)

(iii) PrlC(O)CH₂CH₂CH₂OOCCH₂—(R)Cgl-Aze-Pab(Z)

The title compound was prepared according to the procedure described inExample 1(iii) from H—(R)Cgl-Aze-Pab-Z (6 g; 10.4 mmol) andPrlC(O)CH₂CH₂CH₂OOCCH₂Br (3.5 g; 12.4 mmol; from step (ii) above). Thecrude product was purified by flash chromatography usingheptane:EtOAc:isopropanol (1:2:2) as eluent to give 4.2 g which was thenpurified by using preparative RPLC using 44% acetonitrile in 0.1M NH₄OAcas eluent to give 2.64 g (36%) of the product as a white solid.

¹H-NMR (500 MHz, CDCl₃): δ 9.80-9.22 (b s, 1H), 8.36 (t, 1H), 7.96-7.58(m, 3H), 7.45 (d, 2H), 7.37-7.22 (m, 5H), 5.20 (s, 2H), 4.95-4.88 (dd,1H), 4.72-4.29 (m, 2H), 4.15-4.04 (m, 2H), 4.04-3.88 (m, 2H), 3.40 (t,2H), 3.34 (t, 2H), 3.28-3.17 (m, 2H), 2.85 (d, 1H), 2.67-2.48 (m, 1H),2.23 (t, 2H), 2.14-0.93 (m, 18H). ¹³C-NMR (125 MHz, CDCl₃) amidine andcarbonyl signals: δ 175.3, 172.4, 170.9, 170.4, 168.2, 164.6 FAB-MS:(m+1)=703 (m/z)

EXAMPLE 11 ChNHC(O)CH₂OOCCH₂—(R)Cgl-Aze-Pab-Z

(i) ChNHC(O)CH₂OH

A mixture of cyclohexylamine (9.9 g; 99.8 mmol) and2,5-dioxo-1,4-dioxane (3.0 g, 25.9 mmol) was stirred at 100° C. for 2.5h. The product was concentrated to give 8.1 g (100%) of the product as abrown solid.

¹H-NMR (500 MHz, MeOH-d4): δ 3.92 (s, 2H), 3.75-3.65 (m, 1H), 1.90-1.58(m, 5H), 1.43-1.07 (m, 5H). The signal of two of the protons areobscured by the CD₃OH-signal. ¹³C-NMR (125 MHz, MeOH-d4) amidine andcarbonyl signals: δ 174.0, 62.5, 33.7, 26.5, 26.1, 26.0 The signal ofone of the carbons is obscured by the CD₃OD-signal.

(ii) ChNHC(O)CH₂OOCCH₂Br

To a mixture of ChNHC(O)CH₂OH (8.0 g; 50.9 mmol; from step (i) above)and DMAP (6.2 g; 50.9 mmol) in methylene chloride (80 mL) at 0° C. wasadded dropwise bromoacetyl bromide (4.0 mL; 45.8 mmol). After stirringat room temperature for 1.5 h further portions of bromoacetyl bromide(1.0 mL, 11.4 mmol) and DMAP (1.4 g, 11.4 mmol) were added and thereaction mixture was refluxed for 1.5 h. Water was added and the aqueousphase was extracted with three portions of methylene chloride. Theorganic phase was washed with water, dried with Na₂SO₄, and concentratedto give 10.3 g (73%) of the product as a brown solid.

¹H NMR (400 MHz, CDCl₃): δ 6.12-6.00 (bs, 1H), 4.62 (s, 2H), 3.90 (s,2H), 3.84-3.76 (m, 1H), 1.95-1.86 (m, 2H), 1.75-1.65 (m, 2H), 1.65-1.56(m, 1H), 1.43-1.29 (m, 2H), 1.24-1.10 (m, 3H).

(iii) ChNHC(O)CH₂OOCCH₂—(R)Cgl-Aze-Pab-Z

The title compound was prepared analogously to the procedure describedin Example 1(iii) with starting from H—(R)Cgl-Aze-Pab-Z (6 g; 10.4 mmol)and ChNHC(O)CH₂OOCCH₂Br (3.5 g; 12.4 mmol; from step (ii) above). Thecrude product was purified by flash chromatography usingheptane:EtOAc:isopropanol (5:2:2) as eluent followed by concentrationand then by preparative RPLC using 50% acetonitrile in 0.1M NH₄OAc aseluent. Concentration and freeze drying gave 2.6 g (36%) of the productas a white solid.

¹H-NMR (500 MHz, CDCl₃): δ 9.78-9.25 (bs, 1H), 7.90 (t, 1H), 7.78 (d,2H), 7.44 (d, 2H), 7.38-7.24 (m, 5H), 6.66 (t, 1H), 5.20 (s, 2H),4.90-4.83 (dd, 1H), 4.60-4.45 (m, 2H), 4.18-3.93 (m, 4H), 3.73-3.62 (m,1H), (d, 1H), 3.23, 3.44 (AB, 2H), 2.87, 2.65-2.08 (m, 3H), 1.98-0.93(m, 22H) ¹³C-NMR (125 MHz, CDCl₃) amidine and carbonyl signals: δ 175.1,171.7, 170.7, 168.8, 166.1, 164.4 FAB-MS: (m+1)=703 (m/z)

EXAMPLE 12 (nPr)₂NC(O)CH₂OOCCH₂—(R)Cgl-Aze-Pab-COOCH₂OOCC(CH₃)₃

(i) (nPr)₂NC(O)CH₂OH

A mixture of 2,5-dioxo-1,4-dioxane (2.02 g; 17.4 mmol) anddi-n-propylamine (5 ml; 36.5 mmol) was heated at 50° C. for 1 h and at90° C. for 66 h. Toluene was added and subsequently removed in vacuowith excess di-n-propylamine. The residue was purified by flashchromatography using 10% methanol in methylene chloride as eluent togive 4.18 g (66%) of the desired compound.

¹H NMR (300 MHz, CDCl₃): δ 4.1 (d, 2H), 3.65 (t, 1H), 3.25-3.35 (m, 2H),2.9-3.0 (m, 2H), 1.45-1.6 (m, 4H), 0.8-0.95 (m, 6H)

(ii) (nPr)₂NC(O)CH₂OOCCH—Br

A mixture of (nPr)₂NC(O)CH₂OH (0.743 g; 4.7 mmol; from step (i) above),DCC (0.951 g, 4.6 mmol), and bromoacetic acid (0.704 g; 5.1 mmol) inmethylene chloride (15 ml) was stirred at room temperature for 1.5 h.The precipitate was removed by filtration and the solvent was removedfrom the filtrate in vacuo. Kugelrohr distillation of the residue gave0.66 g (50%) of the desired compound.

¹H NMR (300 MHz, CDCl3): δ 4.8 (s, 2H), 4.0 (s, 2H), 3.2-3.3 (m, 2H),3.05-3.15 (m, 2H), 1.5-1.7 (m, 4H), 0.8-1.0 (dt, 6H)

(iii) Pivaloyloxymethyl 4-nitrophenyl Carbonate

A mixture of silver pivalate (7.5 g; 25 mmol) and iodomethyl4-nitrophenyl carbonate (Alexander et al, J. Med. Chem. (1988) 31, 318;7.99 g; 25 mmol) was refluxed in benzene (50 ml) for 2 h. The benzenewas removed in vacuo and the residue was dissolved in toluene.Filtration through hyflo and purification by flash chromatography usingtoluene as eluent afforded 4.00 g (54%) of the sub-title compound.

¹H NMR (300 MHz; CDCl₃): δ 8.25 (d, 2H), 7.40 (d, 2H), 5.85 (s, 2H), 1.2(s, 2H) ¹³C NMR (75 MHz, CDCl₃) amidine and carbonyl signals: δ 176.77,155.06

(iv) Boc-(R)Cgl-Aze-Pab-COOCH₂OOCC(CH₃)₃

A solution of pivalyloxymethyl 4-nitrophenyl carbonate (1.18 g; 4 mmol;from step (ii) above) in methylene chloride (20 ml) was added at roomtemperature to a solution of Boc-(R)Cgl-Aze-Pab-H (1.88 g; 4 mmol) andtriethylamine (0.66 ml; 4.75 mmol) in methylene chloride (20 ml). After1 h the methylene chloride was replaced by EtOAc and the mixture waspurified by flash chromatography using EtOAc as eluent to give 1.27 g(50%) of sub-title compound.

¹H NMR (300 MHz, CDCl₃): δ 9.5 (bs, 1H), 8.25 (t, 1H), 7.8 (d, 2H), 7.3(d, 2H), 7.0 (bs, 1H), 5.0-4.8 (m, 2H), 4.65-4.5 (m, 1H), 4.5-4.3 (m,2H), 4.2-4.05 (m, 1H), 3.75 (t, 1H), 2.7-2.4 (m, 2H), 1.9-1.45 (m, 5H),1.45-0.8 (m, 24H)

(v) H—(R)Cgl-Aze-Pab-COOCH₂OOCC(CH₃)₃

Boc-(R)Cgl-Aze-Pab-COOCH₂OOCC(CH₃)₃ (327 mg; 0.52 mmol; from step (iv)above) was dissolved in a mixture of methylene chloride (5 ml) and TFA(1.2 ml). After 2 h the reaction mixture was concentrated in vacuo,acetonitrile was added, and the solvent was again removed in vacuo togive crude sub-title product which was used without further purificationin the next step.

(vi) (nPr)₂NC(O)CH₂OOCCH₂—(R)Cgl-Aze-Pab-COOCH₂OOCC(CH₃)₃

The residue from step (v) above was mixed with (nPr)₂NC(O)CH₂OOCCH₂Br(150 mg; 0.53 mmol; from step (ii) above) and K₂CO₃ (480 mg; 3.5 mmol)in THF (5 ml) and heated for 3 h at 40° C. The reaction mixture wasfiltered and concentrated to a crude product which was purified bypreparative RPLC to give 78 mg (21%) of the title compound.

¹H NMR (300 MHz, CDCl₃): δ 9.3-9.6 (bs, 1H), 8.5 (m, 1H), 7.95-8.15 bs,1H), 7.85-7.95 (d, 2H), 7.2-7.3 (d, 2H), 5.8 (s, 2H), 4.8-4.9 (dd, 1H),4.5-4.7 (m, 3H), 4.0-4.4 (m, 3H), 2.8-3.4 (m, 5H), 2.2-2.7 (m, 3H),1.75-1.3 (m, 9H), 1.3-1.0 (m, 14H), 1.0-0.7 (m, 7H). ¹³C NMR (75 MHz,CDCl₃) amidine and carbonyl signals: δ 177.24, 175.30, 171.85, 170.79,168.78, 165.82, 163.14.

EXAMPLE 13 EtOOCCH₂—(R)Cgl-Aze-Pab-COOCH₂OOCC(CH₃)₃

The title compound was prepared analogously to to the proceduredescribed in Example 12(vi) above from crudeBoc-(R)Cgl-Aze-Pab-COOCH₂OOCC(CH₃)₃ (0.41 g; 0.65 mmol; see Example12(iv) above) using acetonitrile (10 ml) as solvent. After stirring overnight at room temperature, the solvent was removed in vacuo and theresidue was partitioned between EtOAc and water. The aqueous phase wasextracted three times with EtOAc and the combined organic phases weredried (Na₂SO₄) and the solvent was removed in vacua. The residue wassubjected to flash chromatography using methylene chloride/methanol aseluent. Freeze drying from glacial acetic acid gave 84 mg (21%) of thetitle compound.

¹H NMR (300 MHz, CDCl₃): δ 9.9 (bs, 1H), 8.5 (t, 1H), 7.35 (d, 2H), 5.85(s, 2H), 5.90 (dd, 2H), 4.6-4.35 (m, 2H), 4.15-4.0 (m, 4), 3.2 (s, 2H),2.85 (d, 1H), 2.7-2.45 (m, 2), 2.0-1.9 (m, 2H), 1.8-1.45 (m, 5H),1.3-0.9 (m, 18H). ¹³C NMR (75 MHz, CDCl₃) amidine and carbonyl signals:δ 177.23, 175.48, 172.29, 170.80, 168.85, 163.14.

EXAMPLE 14 EtOOCCH₂—(R)Cgl-Aze-Pab-COOCH(CH₃)OOCCH₃

(i) Boc-(R)Cgl-Aze-Pab-COOCH(CH₃)OOCCH₃

A solution of Boc-(R)Cgl-Aze-Pab-H (6.38 g; 13.5 mmol), 1-acetoxyethyl4-nitrophenyl carbonate (Alexander et al, J. Med. Chem. (1988) 31, 318)(3.05 g; 12 mmol), and triethylamine (1.95 ml; 14 mmol) in methylenechloride (40 ml) was stirred at room temperature for 16 h followed byaddition of EtOAc. The resulting solution was slightly concentrated andwashed with aqueous Na₂CO₃ (10%), concentrated to a crude product, whichwas purified by flash chromatography using EtOAc as eluent, to give 5.59g (77%) of the sub-title compound.

¹H NMR (300 MHz, CDCl₃): δ 9.5 (bs, 1H), 8.25 (t, 1H), 7.85 (d, 2H),7.35 (d, 2H), 6.95 (q, 1H), 6.7 (bs, 1H), 5.0-4.85 (m, 2H), 4.65-4.5 (m,1H), 4.5-4.25 (m, 2H), 4.2-4.05 (m, 1H) 3.75 (t, 1H), 2.65-2.45 (m, 2H)2.05 (s, 3H), 1.9-1.45 (m, 11H), 1.45-0.8 (m, 12H). ¹³C NMR (75 MHz,CDCl3) amidine and carbonyl signals: δ 172.61, 170.80, 169.54, 168.91,162.50, 156.02.

(ii) H—(R)Cgl-Aze-Pab-COOCH(CH₃)OOCCH₃

The crude sub-title compound was prepared according to the proceduredescribed in Example 12(v) above fromBoc-(R)Cgl-Aze-Pab-COOCH(CH₃)OOCCH₃ (2.21 g; 3.68 mmol; from step (i)above).

(iii) EtOOCCH₂-(R)Cgl-Aze-Pab-COOCH(CH₃)OOCCH₃

The crude H—(R)Cgl-Aze-Pab-COOCH(CH₃)OOCCH₃ from step (ii) above wasdissolved in methylene chloride (150 mL). The mixture was washed with a10% Na₂CO₃ solution and the organic phase was dried with K₂CO₃ andfiltered. To the resulting solution as added K₂CO₃ (756 mg, 5.5 mmol)and ethyl (O-trifluoromethanesulphonyl)-glycolate (790 mg; 3.3 mmol) inmethylene chloride (5 ml). The reaction mixture was stirred for 5-10minutes at room temperature and then concentrated in vacuo. The residuewas dissolved in EtOAc and the resulting mixture was filtered throughcelite. The filtrate was subjected to flash chromatography using EtOAcas eluent followed by HPLC to give 475 mg (22%) of the title compound.

¹NMR (300 MHz, CDCl3): δ 9.5 (bs, 1H), 8.3 (t, 1H), 7.7 (d, 2H), 7.2 (d,2H), 6.85 (q, 1H), 4.8 (t, 1H), 4.45-4.25 (m, 2H), 4.1-3.85 (m, 4H), 3.1(s, 2H), 2.75 (s, 1H), 2.5-2.3 (m, 2H), 1.95 (s, 3H), 1.9-1.8 (m, 1H),1.7-1.25 (m, 8H), 1.25-1.75 (m, 8H). ¹³C NMR (75.5 MHz, CDCl₃) amidineand carbonyl signals: δ 175.26, 172.34, 170.81, 169.49, 168.80, 162.43.

EXAMPLE 15 MeOOCCH₂—(R)Cgl-Aze-Pab-OOCPh

(i) Boc-(R)Cgl-Aze-Pab-OOCPh

To a solution of Boc-(R)Cgl-Aze-Pab-H (1.0 g; 2.1 mmol) and Na₂HPO₄(18.7 g; 105 mmol) in THF (45 mL) at 20° C. was added dropwise dibenzoylperoxide (556 mg; 2.3 mmol) dissolved in THF (10 mL) over 45 minutes.After stirring at 20° C. for 24 h, the reaction mixture was concentratedand the resulting crude product was subjected to preparative RPLC. Thisgave 124 mg (10%) of the sub-title compound as a white solid.

¹H-NMR (500 MHz, CDCl₃): δ 8.26 (m, 1H), 8.09 (m, 2H), 7.72 (m, 2H),7.59 (m, 1H), 7.48 (m, 2H), 7.36 (d, 2H), 5.13 (s, 2H), 4.87-4.98 (m,2H), 4.54-4.61 (m, 1H), 4.33-4.47 (m, 2H), 4.13-4.19 (m, 1H), 3.81 (t,1H), 2.53-2.63 (m, 2H), 1.73-1.86 (m, 3H), 1.66-1.72 (m, 2H), 1.36 (s,9H), 0.968-1.28 (m, 6H). ¹³C-NMR (100 MHz, CDCl₃) amidine and carbonylsignals: δ 172.7, 170.6, 163.9, 157.0, 155.9. LC-MS: m/z 592 (M+H⁺); m/z614 (M+Na⁺).

(ii) H—(R)Cgl-Aze-Pab-OOCPh

To a solution of Boc-(R)Cgl-Aze-Pab-OOCPh (600 mg; 1.01 mmol; from step(i) above) in methylene chloride (18 mL) was added TFA (6 mL) at 20° C.After stirring for 14 h, the reaction mixture was concentrated and theresulting crude product was partitioned between EtOAc:0.1 M NaOH. Thephases were separated and the organic layer was dried (Na₂SO₄) andevaporated. Yield: 480 mg (96%) as a white solid.

¹H-NMR (400 MHz, MeOH-d4): δ 8.18 (m, 2H), 7.77 (m, 2H), 7.64 (m, 1H),7.52 (m, 2H), 7.43 (d, 2H), 4.75-4.81 (m, 1H), 4.50 (s, 2H), 4.18-4.34(m, 2H), 3.12 (d, 1H), 2.57-2.68 (m, 1H), 2.23-2.33 (m, 1H), 1.88-1.96(m, 1H), 1.73-1.84 (m, 2H), 1.59-1.71 (m, 2H), 1.45-1.57 (m, 1H),0.80-1.34 (m, 5H) LC-MS: m/z 492 (M+H⁺); m/z 514 (M+Na⁺)

(iii) MeOOCCH₂—(R)Cgl-Aze-Pab-OOCPh

To a solution of H—(R)Cgl-Aze-Pab-OOCPh (480 mg; 0.97 mmol; from step(ii) above), K₂CO₃ (270 mg; 2 mmol) in acetonitrile (5 mL) at 20° C. wasadded methyl bromoacetate (177 mg; 1.16 mmol). The reaction was stirredat 20° C. for 14 h. The reaction mixture was filtered and concentratedto give a crude product which was purified by preparative RPLC to givegave 269 mg (49%) of the title compound as a white solid.

¹H-NMR (500 MHz, CDCl₃): δ 8.43 (m, 1H, NH), 8.09 (m, 2H), 7.69 (m, 2H),7.59 (m, 1H), 7.47 (m, 2H), 7.34 (m, 2H), 5.27 (s, 2H), 4.93 (dd, 1H),4.59 (dd, 1H), 4.40 (dd, 1H), 4.12 (m, 2H), 3.65 (s, 3H), 2.87 (d, 1H),2.72-2.63 (m, 1H), 2.55-2.48 (m, 1H), 1.96 (m, 1H), 1.74 (m, 2H), 1.67(d, 1H), 1.59 (d, 1H), 1.56-1.50 (m, 1H), 1.29-1.08 (m, 4H), 1.04-0.94(m, 1H) ¹³C-NMR (100 MHz, CDCl₃) amidine and carbonyl signals: δ 175.1,172.5, 170.6, 164.0, 157.1 LC-MS: m/z 564 (M+H⁺)

EXAMPLE 16 MeOOCCH₂—(R)Cgl-Aze-Pab-OH

To a solution of MeOOCCH₂—(R)Cgl-Aze-Pab-OC(O)Ph (260 mg; 0.46 mmol; seeExample 15(iii) above) in THF (4.6 mL) was added KOMe (1.6 mL; 0.29M;0.46 mmol) at 20° C. After 15 minutes of stirring the mixture wasconcentrated and subjected to preparative RPLC. This gave 109 mg (52%)of the title compound as a white solid.

¹H-NMR (500 MHz, MeOH-d4): δ 7.59 (d, 2H), 7.34 (d, 2H), 4.83 (s, 2H),4.82-4.76 (m, 1H), 4.48 (d, 1H), 4.33 (d, 1H), 4.15-4.30 (m, 2H), 3.64(s, 3H), 3.04 (d, 1H), 2.57 (m, 1H), 2.26 (m, 1H), 1.95 (m, 1H), 1.75(m, 2H), 1.58-1.70 (m, 2H), 1.53 (m, 1H), 1.31-1.10 (m, 4H), 1.04 (m,1H) ¹³C-NMR (100 MHz, MeOH-d4): amidine and carbonyl signals: δ 175.9,174.3, 172.7, 155.2 LC-MS: m/z 460 (M+H⁺), m/z 482 (M+Na⁺)

EXAMPLE 17 EtOOCCH₂—(R)Cgl-Aze-Pab-OH

To a solution of EtOOCCH₂—(R)Cgl-Aze-Pab-C(O)OCH(CH₃)OOCCH₃ ₍184 mg;0.31 mmol; see Example 14(iii) above), hydroxylamine hydrochloride (120mg; 1.72 mmol) and triethylamine (0.8 ml; 5.7 mmol) in EtOH (95%; 4.0mL) was added, and the mixture stirred at room temperature for 4 days.The reaction mixture was concentrated and the crude product subjected topreparative RPLC. This gave 85 mg (58%) of the title compound.

¹H-NMR (300 MHz, CD₃OD): δ 7.6 (d, 2H), 7.35 (d, 2H), 4.75-4.85 (m, 1H),4.4-4.55 (m, 2H), 4.0-4.35 (m, 4H), 3.35 (d, 2H), 3.05 (d, 1H), 2.5-2.65(m, 1H), 2.2-2.35 (m, 1H), 1.9-2.05 (m, 1H), 1.4-1.85 (m, 5H), 0.85-1.35(m, 8H) ¹³C-NMR (75.5 MHz, CD₃OD): amidine and carbonyl signals: δ175.97, 173.91, 172.72, 155.23 LC-MS:(m+1)=474 (m/z)

EXAMPLE 18 BnOOCCH₂—(R)Cgl-Aze-Pab-OH

To a solution of hydroxylamine hydrochloride (320 mg; 4.59 mmol) andtriethylamine (1.7 ml; 12.24 mmol) in EtOH, BnOOCCH₂—(R)Cgl-Aze-Pab-Z(1.0 g; 1.52 mmol) was added. The reaction mixture was stirred at roomtemperature for 40 hours and then concentrated. The crude product waspurified by preparative RPLC using 50% acetonitrile in 0.1M NH₄OAc aseluent to give 0.34 g (42%) of the title compound.

LC-MS:(m+1)=536 (m/z)

EXAMPLE 19 nPrOOCCH₂—(R)Cgl-Aze-Pab-Z

The title compound was prepared analogously to the procedure describedin Example 1(iii) from H—(R)Cgl-Aze-Pab-Z×2HCl (700 mg; 1.2 mmol) andn-propyl bromoacetate (268 mg; 1.45 mmol). Yield 259 mg (35%).

FAB-MS:(m+1)=606 (m/z)

EXAMPLE 20 nPrOOCCH₂—(R)Cgl-Aze-Pab-OH

The title compound was prepared analogously to the procedure describedin Example 18 from nPrOOCCH₂—(R)Cgl-Aze-PabZ (182 mg; 0.3 mmol; seeExample 19 above). The crude product was purified by preparative RPLCusing 40% acetonitrile in 0.1M NH₄OAc as eluent to give 74 mg (51%) ofthe desired compound.

LC-MS:(m+1)=488 (m/z)

EXAMPLE 21 iPrOOCCH₂—(R)Cgl-Aze-Pab-OH

The title compound was prepared analogously to the procedure describedin Example 18 from iPrOOCCH₂—(R)Cgl-Aze-Pab-Z (590 mg; 0.7 mmol; seeExample 39 below). Yield 110 mg (32%)

LC-MS:(m+1)=488 (m/z)

EXAMPLE 22 tBuOOCCH₂—(R)Cgl-Aze-Pab-OH

The title compound was prepared analogously to the procedure describedin Example 18 from tBuOOCCH₂—(R)Cgl-Aze-Pab-Z (738 mg; 1.2 mmol; seeExample 37 below). Yield 290 mg (48%).

LC-MS:(m+1)=502 (m/z)

EXAMPLE 23 (nPr)₂NCOCH₂OOCCH₂—(R)Cgl-Aze-Pab(OH)

(i) HOOCCH₂—(R)Cgl(Boc)-Aze-Pab-O-Boc

A solution of HOOCCH₂—(R)Cgl-Aze-Pab-OH (670 mg; 1.5 mmol; see Example28 below), (Boc)₂O (654 mg; 3 mmol), and DMAP (92 mg; 0.75 mmol) inTHF:water (10:1) was stirred at room temperature for 2 h. The reactionmixture was concentrated and purified by preparative RPLC. Freeze dryingyielded 112 mg (12%) of the sub-title compound as a white solid.

LC-MS:(m−1)=643 (m/z)

(ii) (nPr)₂NCOCH₂OOCCH₂—(R)Cgl(Boc)-Aze-Pab-O-Boc

A solution of HOOCCH₂—(R)Cgl(Boc)-Aze-Pab-O-Boc (100 mg; 0.15 mmol; fromstep (i) above), (nPr)₂NCOCH₂OH (27 mg; 0.17 mmol; see Example 12(i)above), EDC (40 mg; 0.21 mmol) and DMAP (10 mg; 0.075 mmol) inacetonitrile (5 mL) was stirred at room temperature for 4 days. Thereaction mixture was concentrated, purified by preparative RPLC andfreeze dried to give 21 mg (18%) of the sub-title compound.

LC-MS:(m−1)=787 (m/z)

(iii) (nPr)₂NCOCH₂OOCCH₂—(R)Cgl-Aze-Pab-OH

A solution of (nPr)₂NCOCH₂—(R)Cgl(Boc)-Aze-Pab-O-Boc (20 mg; 0.025 mmol)in TFA:methylene chloride (1:1) was stirred at room temperature for 5minutes. The reaction mixture was concentrated and freeze dried fromacetonitrile and water to give 5 mg (34%) of the title compound.

LC-MS:(m+1)=587 (m/z)

EXAMPLE 24 ChNHCOCH₂OOCCH₂—(R)Cgl-Aze-Pab-OH

The title compound was prepared analogously to the procedure describedin Example 18 from ChNHCOCH₂OOCCH₂—(R)Cgl-Aze-Pab-Z (118 mg; 0.17 mmol;see Example 11(iii) above). Yield 1.8 mg.

LC-MS:(m+1)=585 (m/z)

EXAMPLE 25 MeNHCOCH₂OOCCCH₂—(R)Cgl-Aze-Pab-OH

The title compound was prepared analogously to the procedure describedin Example 18 from MeNHCOCH₂—(R)Cgl-Aze-Pab-Z (81 mg; 0.12 mmol, seeExample 36 below). Yield 10 mg (16%).

LC-MS:(m+1)=517 (m/z)

EXAMPLE 26 EtOOCCH₂—(R)Cgl-Aze-Pab-OAc

(i) H—(R)Cgl-Aze-Pab-OAc

The sub-title compound was prepared analogously to the method describedin Example 27 below (steps (i), (ii) and (iii)) using acetic acidanhydride instead of propanoic acid anhydride.

LC-MS:(m+1)=430 (m/z)

(ii) EtOOCCH₂—(R)Cgl-Aze-Pab-OAc

The title compound was prepared analogously to the procedure describedin Example 1(iii) above from H—(R)Cgl-Aze-Pab-OAc (370 mg; 0.6 mmol) andethyl bromoacetate (105 mg; 0.63 mmol). Yield 67 mg (22%).

LC-MS:(m+1)=516 (m/z)

EXAMPLE 27 EtOOCCH₂—(R)Cgl-Aze-Pab-OC(O)Et

(i) Boc-(R)Cgl-Az-Pab-OH

To a solution of hydroxylamine hydrochloride and triethylanine in EtOHwas added Boc-(R)Cgl-Aze-Pab-Z (1.0 g; 1.52 mmol). The reaction mixturewas stirred at room temperature for 40 hours and then concentrated. Thecrude product was purified by preparative RPLC.

LC-MS: (m+1)=488 m/z

(ii) Boc-(R)Cgl-Aze-Pab-OC(O)Et

A solution of Boc-(R)Cgl-Aze-Pab-OH (500 mg; 0.91 mmol; from step (i)above) and propanoic acid anhydride (3.5 mL) was stirred at roomtemperature for 45 minutes and then concentrated. The crude product waspurified by preparative RPLC using 50% acetonitrile in 0.1M NH₄OAc aseluent to give 266 mg (54%) of the sub-title compound.

LC-MS: (m+1)=544(m/z)

(iii) H—(R)Cgl-Aze-Pab-OC(O)Et

The sub-title compound was prepared analogously to the proceduredescribed in Example 1(ii) from Boc-(R)Cgl-Aze-Pab-OC(O)Et (238 mg; 0.44mmol; from step (ii) above). Yield 290 mg (100%).

LC-MS:(m+1)=444(m/z)

(iv) EtOOCCH₂—(R)Cgl-Aze-Pab-OC(O)Et

To a solution of H—(R)Cgl-Aze-Pab-OOCEt (300 mg; 0.45 mmol; from step(iii) above) and K₂CO₃ (308 mg; 2.23 mmol) in methylene chloride (6 mL)at 0° C. was added dropwise EtOOCCH₂OSO₂CF₃ (105 mg; 0.45 mmol, preparedfrom triflic anhydride and ethyl glycolate). After the reaction mixturewas stirred at room temperature for 1 h the reaction mixture was washedwith water, citric acid and water, dried (Na₂SO₄) and concentrated. Thecrude product was purified by preparative RPLC using 45% acetonitrile in0.1M NH₄OAc as eluent to give 63 mg (27%) of the title compound.

LC-MS:(m+1)=530(m/z)

EXAMPLE 28 HOOCCH₂—(R)Cgl-Aze-Pab-OH

(i) tBuOOCCH₂—(R)Cgl-Aze-Pab-OOCPh

The sub-title compound was prepared analogously to the proceduredescribed in Example 1(iii) from H—(R)Cgl-Aze-Pab-OOCPh (250 mg; 0.5mmol; see Example 15(ii) above) and t-butylbromoacetate (119 mg; 0.6mmol). Yield 211 mg (69%).

LC-MS:(m+1)=606 (m/z)

(ii) HOOCCH₂—(R)Cgl-Aze-Pab-OOCPh

The sub-title compound was prepared analogously to the proceduredescribed in Example 1(ii) from tBuOOCCH₂—(R)Cgl-Aze-Pab-OOCPh (233 mg;0.3 mmol; from step (i) above). Yield 65 mg (37%).

LC-MS:(m+1)=550 (m/z)

(iii) HOOCCH₂—(R)Cgl-Aze-Pab-OH

A solution of HOOCCH₂—(R)Cgl-Aze-Pab-OOCPh (60 mg; 0.1 mmol; from step(ii) above) and KOMe (0.2M; 0.2 mmol) in THF (10 mL) and methanol (1.5mL) was stirred at room temperature for 5 minutes. The reaction mixturewas concentrated and freeze dried from water and acetonitrile to give 28mg (63%) of the title compound.

LC-MS:(m+1)=446(m/z)

EXAMPLE 29 HOOCCH₂—(R)Cgl-Aze-Pab-O-cis-Oleyl

(i) tBuOOCCH₂—(R)Cgl(Boc)-Aze-Pab-Z

A solution of tBuOOCCH₂—(R)Cgl-Aze-Pab-Z (1.7 g, 2.8 mmol; see Example37 below), (Boc)₂O (672 mg; 3.08 mmol) and DMAP (68 mg; 0.56 mmol) inTHF (30 mL) was stirred at room temperature for 24 h. Additional (Boc)₂O(305 mg; 1.4 mmol) was added at 5° C. After another 24 h the reactionmixture was concentrated and purified by preparative RPLC to give 587 mg(30%) of the desired compound.

EC-MS:(m+1)=720 (m/z)

(ii) tBuOOCCH₂—(R)Cgl(Boc)-Aze-Pab-OH

The sub-title compound was prepared analogously to the proceduredescribed in Example 18 from tBuOOCCH₂—(R)Cgl(Boc)-Aze-Pab-Z (580 mg;0.8 mmol; from step (i) above). Yield 341 mg (71%).

EC-MS:(m+1)=602 (m/z)

(iii) tBuOOCCH₂—(R)Cgl(Boc)-Aze-Pab-O-cis-Oleyl

A solution of tBuOOCCH₂—(R)Cgl(Boc)-Aze-Pab-OH (340 mg; 0.56 mmol; fromstep (ii) above), cis-oleylchloride (170 mg; 0.56 mmol) andtriethylamine (62 mg; 0.61 mmol) in methylene chloride was stirred for 5minutes. The reaction mixture was concentrated and purified bypreparative RPLC to give 326 mg (67%) of the sub-title compound.

EC-MS:(m+1)=867(m/z)

(iv) HOOCCH₂—(R)Cgl-Aze-Pab-O-cis-Oleyl

The title compound was prepared analogously to the procedure describedin Example 1(ii) from tBuOOCCH₂—(R)Cgl(Boc)-Aze-Pab-O-cis-Oleyl (223 mg;0.25 mmol; from step (iii) above).

LC-MS:(m+1)=710 (m/z)

EXAMPLE 30 Cyclooctyl-OOCCH₂—(R)Cgl-Aze-Pab-Z

(i) Cyclooctyl-bromoacetate

Cyclooctanol (1.3 g; 10 mmol) and DMAP (0.3 g) was dissolved inmethylene chloride followed by addition of bromacetyl chloride (1 mL; 12mmol). After stirring for 18 h the reaction mixture was washed withaqueous Na₂CO₃ (2M) and HCl (1M), dried, concentrated and purified byflash chromatography using petroleum ether:methylene chloride (50:50) togive 1.8 g (72%) of the sub-title compound.

(ii) Cyclooctyl-OOCCH₂—(R)Cgl-Aze-Pab-Z

The title compound was prepared analogously to the procedure describedin Example 1(iii) from H—(R)Cgl-Aze-Pab-Z×2HCl (703 mg; 1.2 mmol) andcyclooctyl bromoacetate (363 mg; 1.46 mmol; from step (i) above).

Yield 379 mg (46%). FAB-MS: (m+1)=674(m/z)

EXAMPLE 31 tBuCH₂—OOCCH₂—(R)Cgl-Aze-Pab-Z

The title compound was prepared analogously to the procedure describedin Example 1(iii) from H—(R)Cgl-Aze-Pab-Z×2HCl (2.5 g; 4.3 mmol) andtertbutylmetyl bromoacetate (1.08 g; 5.2 mmol). Yield 1.87 g (69%).

FAB-MS:(m+1)=634 (m/z)

EXAMPLE 32 (2-Me)BnOOCCH₂—(R)Cgl-Aze-Pab-Z

(i) Methylbenzyl Bromoacetate

The sub-title compound was prepared analogously to the proceduredescribed in Example 30(i) from 2-methylbenzylalcohol (5 g; 41 mmol) andbromacetyl chloride (12.6 g; 80 mmol). Yield 8.2 g (82%).

(ii) (2-Me)BnOOCCH₂—(R)Cgl-Aze-Pab-Z

The title compound was prepared analogously to the procedure describedin Example 1 (iii) from H—(R)Cgl-Aze-Pab-Z×2HCl (580 mg; 1 mmol) and2-methylbenzyl bromoacetate (290 mg; 1.2 mmol; from step (i) above).

Yield 30 mg (4.5%). LC-MS:(m+1)=668 (m/z)

EXAMPLE 33 ChCH₂OOCCH₂—(R)Cgl-Aze-Pab-Z

A solution of BnOOCCH₂—(R)Cgl-Aze-Pab-Z (1.41 g; 1.7 mmol) andcyclohexyl methylalcohol (6 mL) in triethylamine (474 μL) and methylenechloride (3 mL) was refluxed for 4 days. The reaction mixture was workedup to give a crude product which was purified by flash chromatographyusing methylene chloride:methanol (95:5) as eluent to give 801 mg (71%)of the title compound.

FAB-MS:(m+1)=660 (m/z)

EXAMPLE 34 ChOOCCH₂—(R)Cgl-Aze-Pab-Z

(i) Cyclohexyl Bromoacetate

The sub-title compound was prepared analogously to the proceduredescribed in Example 32(i) above from cyclohexanol (1 g; 10 mmol) andbromacetylchloride (1 mL; 12 mmol).

(ii) ChOOCCH₂—(R)Cgl-Aze-Pab-Z

The title compound was prepared analogously to the procedure describedin Example 1(iii) from H—(R)Cgl-Aze-Pab-Z×2HCl (2.5 g; 4.32 mmol) andcyclohexyl bromoacetate (1.5 g; 5.2 mmol). Yield 1.7 g (60%).

FAB-MS:(m+1)=646 (m/z)

EXAMPLE 35 PhC(Me)₂OOCCH₂—(R)Cgl-Aze-Pab-Z

(i) 2-Phenyl-2-propyl bromoacetate

The sub-title compound was prepared analogously to the proceduredescribed in Example 30(i) from 2-phenyl-2-propanol (3 g; 22 mmol) andbromacetylchloride (4.16 g, 26 mmol). Yield 1.2 g (44%).

(ii) PhC(Me)₂OOCCH₂—(R)Cgl-Aze-Pab-Z

The title compound was prepared analogously to the procedure describedin Example 1(iii) from H—(R)Cgl-Aze-Pab-Z×2HCl (1.2 g; 2.2 mmol) and2-phenyl-2-propyl bromoacetate (640 mg; 2.5 mmol; from step (i) above).

Yield 1.3 g (86%). ¹H-NMR (500 MHz; CDCl₃) δ 9.3 (br s, 1H), 8.35 (t,1H), 7.75 (d, 2H), 7.45 (d, 2H), 7.30-7.05 (m, 10H or 11H), 5.15 (s,2H), 4.78 (t, 1H), 4.40-4.30 (AB part of ABX spectrum, 2H), 3.95 (q,1H), 3.74 (q, 1H), 3.27-3.19 (AB-spectrum, 2H), 2.72 (d, 1H), 2.43 (q,2H), 1.93 (br d, 1H), 1.75-1.60 (m, 9H or 10H), 1.54 (d, 1H), 1.49-1.40(m, 1H), 1.25-1.0 (m, 4H), 0.92 (q, 1H)

EXAMPLE 36 MeNCOCH₂OOCCH₂—(R)Cgl-Aze-Pab-Z

The title compound was prepared analogously to the procedure describedin Example 1(iii) from H—(R)Cgl-Aze-Pab-Z×2HCl (1.0 g; 1.7 mmol) andMeNHCOCH₂OOCCH₂Br (440 mg; 2 mmol; prepared analogously to theprocedures described in Example 11 above (steps (i), (ii) and (iii))using methylamine instead of cyclohexylamine). Yield 380 mg (35%).

FAB-MS:(m+1)=635 (m/z)

EXAMPLE 37 tBuOOCCH₂—(R)Cgl-Aze-PabZ

The title compound was prepared analogously to the procedure describedin Example 1(iii) from H—(R)Cgl-Aze-Pab-Z×2HCl (500 mg; 1.0 mmol) andt-butyl bromoacetate (231 mg; 1.2 mmol). Yield 420 mg (69%).

LC-MS:(m+1)=620 (m/z)

EXAMPLE 38 (Me)₂CHC(Me)₂OOCCH₂—(R)Cgl-Aze-Pab-Z

The title compound was prepared analogously to the procedure describedin Example 1(iii) from H—(R)Cgl-Aze-PabZ (787 mg; 1.4 mmol) and2,3-dimethyl-2-butyl bromoacetate (364 mg; 1.63 mmol). Yield 590 mg(67%).

FAB-MS:(m+1)=648(m/z)

EXAMPLE 39 iPrOOCCH₂—(R)Cgl-Aze-Pab-Z

The title compound was prepared analogously to the procedure describedin Example 1(iii) from H—(R)Cgl-Aze-Pab-Z (700 mg; 1.2 mmol) andisopropyl bromoacetate (262 mg; 1.5 mmol). Yield 225 mg (31%)

FAB-MS:(m+1)=606(m/z)

EXAMPLE 40 BnOOCCH₂—(R)Cgl-Aze-Pab-COOPh(4-OMe)

(i) Boc-(R)Cgl-Aze-Pab-COOPh(4-OMe)

The sub-title compound was prepared analogously to the proceduredescribed in Example 1(i) from Boc-(R)Cgl-Aze-Pab-H and 4-methoxyphenylchloroformate.

FAB-MS:(m+1)=622(m/z)

(ii) H—(R)Cgl-Aze-Pab-COOPh(4-OMe)×2HCl

The sub-title compound was prepared analogously to the proceduredescribed in Example 4(ii) from Boc-(R)Cgl-Aze-Pab-COOPh(4-OMe) (fromstep (i) above).

(iii) BnOOCCH₂—(R)Cgl-Aze-Pab-COOPh(4-OMe)

The title compound was prepared analogously to the procedure describedin Example 1(iii) from H—(R)Cgl-Aze-Pab-COOPh(4-OMe)×2HCl (85 mg; 0.16mmol; from step (iii) above) and benzyl bromoacetate (90 mg; 0.2 mmol).Yield 60 mg (56%).

FAB-MS:(m+1)=670 (m/z)

EXAMPLE 41 ChCH₂OOCCH₂—(R)Cgl-Aze-Pab-COOPh(4-OMe)

The title compound was prepared analogously to the procedure describedin Example 1(iii) from H—(R)Cgl-Aze-Pab-COOPh(4-OMe) (554 mg; 0.64 mmol;see Example 40(ii) above) and cyclohexylmethyl bromoacetate (165 mg; 0.7mmol). Yield 34 mg (8%).

FAB-MS:(m+1)=676 (m/z)

EXAMPLE 42 (2-Me)BnOOCCH₂—(R)Cgl-Aze-Pab-COOPh(4-OMe)

The title compound was prepared analogously to the procedure describedin Example 1(iii) from H—(R)Cgl-Aze-Pab-COOPh(4-OMe) (522 mg; 1 mmol;see Example 40(ii) above) and 2-(methyl)benzyl bromoacetate (365 mg; 1.5mmol). Yield 158 mg (23%).

LC-MS:(m+1)=684 (m/z)

EXAMPLE 43 EtOOCCH₂—(R)Cgl-Aze-Pab-COOPh(4-Me)

(i) Boc-(R)Cgl-Aze-Pab-COOPh(4-Me)

The sub-title compound was prepared analogously to the proceduredescribed in Example 1(i) from Boc-(R)Cgl-Aze-Pab (1.96 g; 4.56 mmol)and 4-tolyl-chloroformate (850 mg; 4.99 mmol). Yield 1.39 g (55%).

FAB-MS:(m+1)=606(m/z)

(ii) H—(R)Cgl-Aze-Pab-COOPh(4-Me)

The sub-title compound was prepared analogously to the proceduredescribed in Example 4(ii) from Boc-(R)Cgl-Aze-Pab-COOPh(4-Me) (388 mg;0.64 mmol; from step (i) above). Yield 293 mg (91%).

FAB-MS:(m+1)=506(m/z)

(iii) EtOOCCH₂—(R)Cgl-Aze-Pab-COOPh(4-Me)

The title compound was prepared analogously to the procedure describedin Example 1(iii) from H—(R)Cgl-Aze-Pab-COOPh(4-Me) (288 mg; 0.6 mmol;from step (ii) above) and ethyl bromoacetate (114 mg; 0.7 mmol). Yield81 mg (24%).

FAB-MS:(m+1)=592 (m/z)

EXAMPLE 44 BnOOCCH₂—(R)Cgl-Aze-Pab-COOPh(4-Me)

The title compound was prepared analogously to the procedure describedin Example 1(iii) from H—(R)Cgl-Aze-Pab-COOPh(4-Me) (272 mg; 0.54 mmol;see Example 43(ii) above) and benzyl bromoacetate (147 mg; 0.6 mmol).Yield 10⁷ mg (31%).

FAB-MS:(m+1)=654(m/z)

EXAMPLE 45 BnOOCCH₂—(R)Cgl-Aze-Pab-COO-nBu

The title compound was prepared analogously to the procedure describedin Example 1(iii) from H—(R)Cgl-Aze-Pab-COO-nBu×2HCl (400 mg; 0.74 mmol;see Example 9(ii) above) and benzyl bromoacetate (210 mg; 0.88 mmol).Yield 220 mg (48%).

FAB-MS:(m+1)=620 (m/z)

EXAMPLE 46 iPrOOCCH₂—(R)Cgl-Aze-Pab-COOCH₂CH═CH₂

The title compound was prepared analogously to the procedure describedin Example 1(iii) from H—(R)Cgl-Aze-Pab-COOCH₂CH═CH₂×2TFA (456 mg; 0.67mmol; see Example 1(ii) above) and isopropyl bromoacetate (145 mg; 0.8mmol). Yield 294 mg (79%).

FAB-MS:(m+1)=556 (m/z)

EXAMPLE 47 EtOOCCH₂—(R)Cgl-Aze-Pab-COO-iBu

(i) Boc-Pab-COO-tBu

To a solution of Boc-Pab-H(500 mg; 2.0 mmol; prepared from Pab-Z and(Boc)₂O (forming Boc-Pab-Z), followed by hydrogenation over Pd/C) andtriethylamine (400 mg; 4.0 mmol) in methylene chloride (10 mL) was addedi-butyl chloroformate (270 mg; 2.2 mmol) at 0° C. After stirring for 5h, water was added. The organic phase was dried (Na₂SO₄) andconcentrated to give 530 mg (76%) of the sub-title compound.

¹H-NMR (500 MHz, CDCl₃) δ 9.5 (bs, 1H), 7.82 (d, 2H), 7.31 (d, 2H), 6.6(bs, 1H), 5.0 (bs, 1H), 4.33 (bd, 2H), 3.93 (d, 2H), 2.04 (m, 1H), 1.45(s, 9H), 0.97 (d, 6H)

(ii) H-Pab-COO-iBu×2HCl

The sub-title compound was prepared analogously to the proceduredescribed in Example 4(ii) from Boc-Pab-COO-iBu (520 mg; 1.5 mmol; fromstep (i) above). Yield 430 mg (88%).

¹H-NMR (500 MHz, MeOD) δ 7.89 (d, 2H), 7.75 (d, 2H), 4.30 (s, 2H), 4.17(d, 2H), 2.11-2.05 (m, 1H), 1.02 (d, 6H)

(iii) Boc-(R)Cgl-Aze-Pab-COO-iBu

To a solution of Boc-(R)Cgl-Aze-OH (480 mg; 1.4 mmol),H-Pab-COO-iBu×2HCl (430 mg; 1.3 mmol; from step (ii) above) and DMAP(650 mg; 5.3 mmol) in acetonitrile (20 mL) was added EDC (270 mg; 1.4mmol). After stirring for 3 days at room temperature the reactionmixture was concentrated and then dissolved in water and EtOAc. Theorganic phase was washed with NaHCO₃ (aq) and dried (Na₂SO₄),concentrated and purified by flash chromatography using EtOAc as eluentto give 510 mg (52%) of the sub-title compound.

(iv) H—(R)Cgl-Aze-Pab-COO-iBu×2HCl

The sub-title compound was prepared analogously to the proceduredescribed in Example 4(ii) from Boc-(R)Cgl-Aze-Pab-COO-iBu (500 mg; 0.88mmol; from step (iii) above). Yield 360 mg (87%).

(v) EtOOCCH₂—(R)Cgl-Aze-Pab-COO-iBu

The title compound was prepared analogously to the procedure describedin Example 1(iii) from H—(R)Cgl-Aze-Pab-COO-iBu×2HCl (290 mg; 0.53 mmol;from step (iv) above) and ethyl bromoacetate (110 mg; 0.64 mmol). Yield140 mg (47%).

FAB-MS:(m+1)=558 (m/z)

EXAMPLE 48 BnOOCCH₂—(R)Cgl-Aze-Pab-COO-nPr

The title compound was prepared analogously to the procedure describedin Example 1(iii) from H—(R)Cgl-Aze-Pab-COO-nPr×2TFA (902 mg; 1.3 mmol;see Example 5(ii) above) and benzyl bromoacetate (362 mg; 1.6 mmol).Yield 199 mg (25;%).

¹H-NMR: (400 MHz; CDCl₃) δ 8.43 (bs, 1H), 7.78 (d, 2H), 7.38-7.27 (m,7H), 5.05 (s, 2H), 4.90 (dd, 1H), 4.56-4.39 (AB part of ABX spectrum,2H), 4.12-4.03 (m, 3H), 3.98-3.91 (q, 1H), 3.33-3.22 (AB-spectrum, 2H),2.85 (d, 1H), 2.65-0.94 (m, 19H)

EXAMPLE 49 EtOOCCH₂—(R)Cgl-Aze-Pab-COOCH₂OOCCh

(i) EtSCOOCH₂OOCCh

To a solution of tetrabutylammonium hydrogensulphate (15.6 g, 45.6 mmol)and cyclohexane carboxylic acid (5.85 g, 46 mmol) in methylene chloridewas added NaOH (9.1 mL, 10M; 68 mmol) at 0° C. After stirring for 5minutes the reaction mixture was filtered, washed with methylenechloride, dissolved in toluene, concentrated and dissolved in THF togive [Bu₄N]⁺[OOCCh]⁻. EtSCOOCH₂Cl (4 g; 25.9 mmol; see Folkmann andLund, J. Synthesis, (1990), 1159) was added to the THF solution of[Bu₄N]⁺[OOCCh]⁻ at room temperature. After stirring at room temperaturefor 12 h the reaction mixture was concentrated and purified by flashchromatography to give 2.57 g (40%) of the sub-title compound.

¹H-NMR (400 MHz, CDCl₃) diagnostic peaks δ 5.80 (s, 2H, O—CH₂—O), 2.85(q, 2H, CH₂—S)

(ii) ClCOOCH₂OOCCh

To EtSCOOCH₂OOCCh (2.9 g; 11.8 mmol; from step (i) above) was addeddropwise SO₂Cl₂ (3.18 g; 23.6 mmol) at 0° C. After stirring for 30minutes the reaction mixture was concentrated to give 1.82 g (70%) ofthe desired compound.

¹H-NMR (500 MHz, CDCl₃) diagnostic peaks δ 5.82 (s, 2H, O—CH₂—O)

(iii) Boc-(R)Cgl-Aze-Pab-COOCH₂OOCCh

The sub-title compound was prepared analogously to the proceduredescribed in Example 1(i) from Boc-(R)Cgl-Aze-Pab-H (750 mg; 1.59 mmol)and ClCOOCH₂OOCCh (460 mg; 2.1 mmol; from step (ii) above). The crudeproduct was purified by preparative RPLC. Yield 355 mg (9%).

FAB-MS:(m+1)=656(m/z)

(iv) H—(R)Cgl-Aze-Pab-COOCH₂OOCCh×2TFA

The sub-title compound was prepared analogously to the proceduredescribed in Example 1(ii) from Boc-(R)Cgl-Aze-Pab-COOCH₂OOCCh (fromstep (iii) above).

(v) EtOOCCH₂—(R)Cgl-Aze-Pab-COOCH₂OOCCh

The title compound was prepared analogously to the procedure describedin Example 1(iii) from H—(R)Cgl-Aze-Pab-COOCH₂OOCCh×2TFA (193 mg; 0.35mmol; from step (iv) above) and ethyl trifluoroacetate (83 mg; 0.35mmol). Yield 87 mg (39%).

¹H—NMR (400 MHz, CDCl₃) δ 8.48 (t br, 1H), 7.83 (d, 2H), 7.37 (d, 2H),5.86 (s, 2H), 4.95 (dd, 1H), 4.15-4.39 (AB part of ABX spectrum, 2H),4.18-4.05 (m, 5H), 3.26-3.17 (AB-spectrum, 2H), 2.87 (d, 1H), 2.75-0.95(m, 29H)

EXAMPLE 50 EtOOCCH₂—(R)Cgl-Aze-Pab-COOCH₂OOCCH₂Ch

The title compound was prepared analogously to the procedure describedis in Example 49 above starting with cyclohexyl acetic acid instead ofcyclohexane carboxylic acid. Yield 74 mg (17%).

FAB-MS:(m+1)=656(m/z)

EXAMPLE 51 EtOOCCH₂—(R)Cgl-Aze-Pab-COOCH(Me)OOCPh

The title compound was prepared analogously to the procedure describedin Example 49 above starting with EtSCOOCH(CH₃)Cl (prepared fromClCOCH(CH₃)Cl and EtSH using the procedure described by Folkmann et alin J. Synthesis, (1990), 1159) instead of EtSCOOCH₂Cl. Yield 70 mg(23%).

FAB-MS:(m+1)=650 (m/z)

EXAMPLE 52 EtOOCCH₂—(R)Cgl-Aze-Pab-COOCH₂OOCPh

The title compound was prepared analogously to the procedure describedin Example 49 above using benzoic acid instead of cyclohexane carboxylicacid. Yield 50 mg (39%).

¹H-NMR (300 MHz, CDCl₃) δ 9.73-9.25 (s br, 1H), 8.45 (t, 1H), 8.05 (d,2H), 7.83 (d, 2H), 7.60-7.10 (m, 6H), 6.10 (s, 2H), 4.96-4.84 (dd, 1H),4.62-4.30 (ABX, 2H), 4.20-3.93 (m, 4H), 3.25 (s, 2H), 2.84 (d, 1H),2.73-2.41 (m, 2H), 2.41-0.87 (m, 15H) ¹³C-NMR (300 MHz, CDCl₃, amidineand carbonyl carbons) δ 163.1, 165.3, 169.0, 170.8, 172.3, 175.5

EXAMPLE 53 BnOOCCH₂—(R)Cgl-Aze-Pab-COOCH(Me)OAc

The title compound was prepared analogously to the procedure describedin Example 1(iii) from H—(R)Cgl-Aze-Pab-COOCH(CH₃)OC(O)CH₃ (108 mg; 0.21mmol; see Example 14(ii) above) and benzyl bromoacetate (36 μL; 0.23mmol). Yield 41 mg (30%).

FAB-MS:(m+1)=650(m/z)

EXAMPLE 54 EtOOCCH₂—(R)Cgl-Aze-Pab-COOCH₂OAc

(i) H—(R)Cgl-Aze-Pab-COOCH₂OAc×2TFA

The sub-title compound was prepared analogously to the proceduredescribed in Example 14 (steps (i) and (ii)) above using acetoxymethyl4-nitrophenylcarbonate (prepared analogously to the method described inExample 12(iii) using silver acetate instead of silver pivalate). Workup gave the sub-title compound which was used in the next step withoutfurther purification.

(ii) EtOOCCH₂—(R)Cgl-Aze-Pab-COOCH₂OAc

The title compound was prepared analogously to the procedure describedin Example 1(iii) above from H—(R)Cgl-Aze-Pab-COOCH₂OAc×2TFA (0.83 mmol;from step (i) above) and ethyl bromoacetate (2.2 mmol). Yield 286 mg.

FAB-MS:(m+1)=574 (m/z)

EXAMPLE 55 tBuOOCCH₂—(R)Cgl-Aze-Pab-COOCH₂OAc

The title compound was prepared analogously to the procedure describedin Example 1(iii) above from H—(R)Cgl-Aze-Pab-COOCH₂OAc×2TFA (0.313mmol; see Example 54(i) above) and t-butyl bromoacetate (73 mg; 0.376mmol). Yield 156 mg (83%).

FAB-MS:(m+1)=602(m/z)

EXAMPLE 56 BnOOCCH₂—(R)Cgl-Aze-Pab-COOCH₂OOC-tBu

The title compound was prepared analogously to the procedure describedin Example 1(iii) from H—(R)Cgl-Aze-Pab-COOCH₂OOC-tBu (379 mg; 0.71mmol; see Example 12(v) above) and benzyl bromoacetate (135 μL; 0.85mmol). Yield 146 mg (30%).

FAB-MS:(m+1)=678(m/z)

EXAMPLE 57 EtOOCCH₂—(R)Cgl-Aze-Pab-COOCH₂CCl₃

(i) Boc-(R)Cgl-Aze-Pab-COOCH₂CCl₃

The sub-title compound was prepared analogously to the proceduredescribed in Example 1(i) from Boc-(R)Cgl-Aze-Pab-H (1.0 g; 2.12 mmol),2M NaOH (11.7 ml) and trichloroethyl chloroformate (494 mg; 2.33 mmol).Yield 1.08 g (79%).

(ii) H—(R)Cgl-Aze-Pab-COOCH₂CCl₃

The sub-title compound was prepared analougously to the proceduredescribed in Example 1(ii) from Boc-(R)Cgl-Aze-Pab-COOCH₂CCl₃ ₍1.04 g;1.607 mmol; from step (i) above). Yield 1.43 g (99%).

¹H-NMR: (500 MHz; CD₃OD) δ 7.79 (d, 2H), 7.61 (d, 2H), 5.10 (s, 2H),4.87-4.81 (m, 2H), 4.63-4.52 (q, 2H), 4.41-4.34 (m, 1H), 4.30-4.24 (m,1H), 3.72 (d, 1H), 2.72-2.63 (m, 1H), 2.32-2.25 (m, 1H), 1.88-1.10 (m,14H)

(iii) EtOOCCH₂-(R)Cgl-Aze Pab-COOCH₂CCl₃

The title compound was prepared analogously to the procedure describedin Example 1(iii) from H—(R)Cgl-Aze-Pab-COOCH₂CCl₃ (400 mg; 0.52 mmol;from step (ii) above) and ethyl bromoacetate (95 mg; 0.57 mmol). Yield 8mg (23%).

¹H-NMR: (500 MHz; CDCl₃) δ 8.47 (bt, 1H), 7.83 (d, 2H), 7.48 (bs, 1H),7.31 (d, 2H), 4.92 (dd, 1H), 4.85 (s, 2H), 4.58-4.39 (AB part of ABXspectrum, 2H), 4.16-4.06 (m, 4H), 3.24 (s, 2H), 4.87 (d, 1H), 2.65-2.59(m, 1H), 2.56-2.48 (m, 1H), 2.10-0.95 (m, 16H)

EXAMPLE 58 MeOOC—C(═CEt)CH₂OOC₂OOCH₂—(R)Cgl-Aze-Pab-Z

(i) MeOOC—C(═CH)C(OH)Et

Propionaldehyde (10.1 g; 0.174 mol) was added dropwise to a solution ofmethyl acrylate (10 g; 0.116 mol) and 1,4-iazobicyclo[2,2,2]octane (1.3g; 0.0116 mol). The reaction mixture was stirred at room temperature for14 days. Ethyl acetate (150 ml) was added. The organic phase was washedwith water and brine, dried (Na₂SO₄), filtered and concentrated to givethe desired compound. Yield 15.5 g (93%).

¹H-NMR: (400 MHz; CDCl₃) δ 6.24 (s, 1H), 5.81 (s, 1H), 4.34 (t, 1H),3.78 (s, 3H), 2.82 (bs, 1H), 1.69 (m, 2H), 0.95 (t, 3H)

(ii) MeOOC—C(═CEt)CH₂Br

HBr (6.5 ml, ˜48%) was added dropwise to MeOOC—C(═CH)C(OH)Et (3 g; 20.8mmol; from step (i) above) at 0° C. After 5 minutes H₂SO₄ (conc.; 6 ml)was added dropwise. The reaction mixture was stirred for 12 hours atroom temperature. Two phases was separated and the top phase was dilutedwith ether. The ether phase was washed with water and aqueous NaHCO₃,dried (Na₂SO₄ and charcoal) and concentrated. The residue was purifiedby flash chromatography. Yield 1.7 g (40%).

¹H-NMR: (400 MHz; CDCl₃) δ 6.97 (t, 3H), 4.23 (s, 2H), 3.8 (s, 3H), 2.32(m, 2H), 1.13 (t, 3H)

(iii) tBuOOCCH₂—(R)Cgl-Aze-Pab-Z

The sub-title compound was prepared analogously to the proceduredescribed in Example 1(iii) from H—(R)Cgl-Aze-Pab-Z (2.1 g; 3.6 mmol)and t-butyl bromoacetate (780 mg, 4.0 mmol). Yield 1.73 g (78%).

(iv) HOOCCH₂—(R)Cgl-Aze-Pab-Z

A solution of tBuOOCCH₂—(R)Cgl-Aze-Pab-Z (from step (ii) above) and TFAin methylene chloride was stirred in room temperature for 3 h. Thereaction mixture was concentrated and freeze dried from water and HCl(conc.; 10 eq.).

(v) MeOOC—C(═CEt)CH—OOCCH₂—(R)Cgl-Aze-Pab-Z

A solution of HOOCCH₂—(R)Cgl-Aze-Pab-Z (263 mg; 0.41 mmol; from step(iv) above), NaOH (1M; 1.239 ml; 1.239 mmol) and water (4 ml) was freezedried. DMF (5 ml) was added, followed by dropwise addition ofMe-OOC—C(═CEt)CH₂Br (103 mg; 0.496 mmol; from step (ii) above) at 0° C.The reaction mixture was stirred for 24 h at room temperature, dilutedwith toluene (5 ml), washed with water, dried (Na₂SO₄) and concentrated.The residue was purified by flash chromatography using EtOAc:methanol(95:5) as eluent. Yield 95 mg (33%).

FAB-MS: (m+1)=690 (m/z)

EXAMPLE 59 MenOOCCH₂—(R)Cgl-Aze-Pab-COOPh(4-OMe)

(i) MenCOCCH₂Br

The sub-title compound was prepared analogously to the proceduredescribed in Example 30(i) above from MenOH (10 mmol) and bromoacetylchloride (12 mmol). Yield 1.5 g (54%).

(ii) MenOOCCH₂—(R)Cgl-Aze-Pab-COOPh(4-OMe)

The title compound was prepared analogously to the procedure describedin Example 1(iii) from H—(R)Cgl-Aze-Pab-COO-Ph(4-OMe) (521 mg; 1 mmol;see Example 40(ii) above) and MenOOCCH₂Br (416 mg; 1.5 mmol; from step(i) above). Yield 36 mg (5%).

FAB-MS: (m+1)=718 (m/z)

EXAMPLE 60 tBuOOCCH₂—(R)Cgl-Aze-Pab-COOnPr

The title compound was prepared analogously to the procedure describedin Example 1(iii) from H—(R)Cgl-Aze-Pab-COO-nPr (575 mg; 0.837 mmol; seeExample 5(ii) above) and t-butyl bromoacetate (196 mg; 1.01 mmol). Yield110 mg (23%).

LC-MS:(m+1)=572 (m/z)

EXAMPLE 61 MenOOCCH₂—(R)Cgl-Aze-Pab-Z

The title compound was prepared analogously to the procedure describedin Example 1(iii) from H—(R)Cgl-Aze-Pab-Z (0.7 g; 1.21 mmol) andMenOOCCH₂Br (0.4 g; 1.45 mmol; see Example 59(i) above). Yield 0.33 g(38%).

FAB-MS:(m+1)=702 (m/z)

EXAMPLE 62 BnOOCCH₂—(R)Cgl-Aze-Pab-COO-Bn(4-NO₂)

(i) Boc-(R)Cgl-Aze-Pab-COO-Bn(4-NO₂)

The sub-title compound was prepared analogously to the proceduredescribed in Example 1(i) from Boc-(R)Cgl-Aze-Pab-H (1.03 g; 2.18 mmol),2M NaOH (24 mL) and 4-NO₂-benzyl chloroformate (518 mg; 2.4 mmol). Yield1.32 g (93%).

FAB-MS: (m+1)=651 (m/z)

(ii) H—(R)Cgl-Aze-Pab-COO-Bn(4-NO₂)

The sub-title compound was prepared analogously to the proceduredescribed in Example 4(ii) from Boc-(R)Cgl-Aze-Pab-COO-Bn(4-NO₂) (1.32mg; 2.03 mmol; from step (i) above). Yield 1.0 g (79%).

FAB-MS:(m+1)=551 (m/z)

(iii) BnOOCCH₂—(R)Cgl-Aze-Pab-COO-Bn(4-NO₂)

The title compound was prepared analogously to the procedure describedin Example 1(iii) from H—(R)Cgl-Aze-Pab-COO-Bn(4-NO₂) (0.5 g; 0.80 mmol;from step (ii) above) and benzyl bromoacetate (220 mg; 0.90 mmol).

FAB-MS:(m+1)=699(m/z)

EXAMPLE 63 EtOOCCH₂—(R)Cgl-Aze-Pab-Bn(4-NO₂)

The title compound was prepared analogously to the procedure describedin Example 1(iii) from H—(R)Cgl-Aze-Pab-COO-Bn(4-NO₂) (211 mg; 0.38mmol; see Example 62(ii) above) and ethyl bromoacetate (47 μl; 0.42mmol). Yield 44 mg (18%).

¹H-NMR: (300 MHz; CDCl3) δ 9.55 (bs, 1H), 8.50 (bt, 1H), 8.20 (d, 2H),7.80 (d, 2H), 7.60 (d, 2H), 7.35 (d, 2H), 6.87 (bs, 1H), 4.95 (dd, 1H),4.65-4.40 (AB part of ABX spectrum, 2H), 4.18-4.04 (m, 5H), 3.27-3.15(AB-spectrum, 2H), 2.87 (d, 1H), 2.75-2.60 (m, 1H), 2.57-2.45 (m, 1H),2.00-0.95 (m, 16H).

EXAMPLE 64 PrlC(O)CH₂OOCCH₂—(R)Cgl-Aze-PabZ

(i) PrlC(O)CH₂OH

A mixture of 2,5-dioxo-1,4-dioxane (2.0 g; 17 mmol) and pyrrolidine (8ml; 97 mmol) was refluxed for 1 h. The excess pyrrolidine was removed byevaporation. Yield 4.4 g (99%).

FAB-MS(m+1)=130(m/z)

(ii) PrlC(O)CH₂OOCCH₂Br

To a solution of PrlC(O)CH₂OH (0.4 g; 3.1 mmol; from step (i) above) inDMF (15 ml) was added dropwise bromoacetyl bromide (0.63 g; 3.1 mmol) at0° C. The reaction mixture was stirred for 1.5 h at 0° C. and 3 h atroom temperature. Additional bromoacetyl bromide (0.63 g; 3.1 mmol) wasadded and the reaction mixture was heated to 80° C., stirred at roomtemperature for 12 h and concentrated. Yield 320 mg (41%)

FAB-MS(m+1)=252(m/z)

(iii) PrlC(O)CH₂OOCCH₂—(R)Cgl-Aze-Pab-Z

The title compound was prepared analogously to the procedure describedin Example 1(iii) from H—(R)Cgl-Aze-Pab-Z (580 mg; 1 mmol) andPrlC(O)CH₂OOCCH₂Br (300 mg; 1.2 mmol; from step (ii) above). Yield 400mg (60%).

FAB-MS(m+1)=675 (m/z)

¹H-NMR: (500 MHz; CDCl₃) δ 9.66-9.42 (bs, 1H), 8.64-8.56 (m, 1H),8.03-7.93 (d, 2H), 7.89-7.66 (bs, 1H), 7.45 (d, 2H), 7.45-7.25 (m, 5H),5.20 (s, 2H), 4.98-4.92 (dd, 1H), 4.82-4.74 (m, 1H), 4.62, 4.58 (ABspectrum, 2H), 4.26-4.05 (m, 3H), 3.47-3.16 (m, 6H), 2.95 (d, 1H),2.78-2.68 (m, 1H), 2.54-2.42 (m, 1H), 2.03-1.95 (m, 16H)

EXAMPLE 65 (2-Me)BnOOCCH₂—(R)Cgl-Aze-Pab-COO-Bn(4-NO₂)

The title compound was prepared analogously to the procedure describedin Example 1(iii) from H—(R)Cgl-Aze-Pab-COO-Bn(4-NO₂) (500 mg; 0.80mmol; see Example 62(ii) above) and 2-(methyl)benzyl bromoacetate (234mg; 0.96 mmol; see Example 32(i) above). Yield 528 mg (92%).

¹H-NMR: (400 MHz, CDCl₃) δ 9.34 (bs, 1H), 8.38 (t, 1H), 8.09 (d, 2H),7.72 (d, 2H), 7.48 (d, 2H), 7.37 (bs, 1H), 7.23 (d, 2H), 7.17-7.05 (m,4H), 5.18 (s, 2H), 5.00 (s, 2H), 4.81 (dd, 1H), 4.45-4.34 (AB part ofABX spectrum, 2H), 4.04-3.97 (q, 1H), 3.93-3.86 (q, 1H), 3.27-3.17 (ABspectrum, 2H), 2.79 (d, 1H), 2.54-2.35 (m, 2H), 2.22 (s, 3H), 1.91-1.84(bd, 1H), 1.71-1.39 (m, 5H), 1.19-0.84 (m, 4H).

EXAMPLE 66 MeOOCCH₂—(R)Cgl-Aze-Pab-COOEt

The title compound was prepared analogously to the procedure describedin Example 1(iii) from H—(R)Cgl-Aze-Pab-COOEt (305 mg; 0.69 mmol; seeExample 4(ii) and methyl bromoacetate (126 mg; 0.83 mmol). Yield 188 mg(53%).

LC-MS:(m+1)=516(m/z)

EXAMPLE 67 (nPr)₂NC(O)CH₂OOCCH₂—(R)Cgl-Aze-Pab-COO-Bn(4-NO₂)

(i) (nPr)₂NC(O)CH₂OOCCH₂Cl

A mixture of (nPr)₂NC(O)CH₂OH (244 mg; 1.53 mmol; see Example 12(i)above) and bromacetyl chloride (270 mg; 1.72 mmol) was stirred at roomtemperature for 12 hours. The mixture was poured into aqueous NaHCO₃ andextracted with methylene chloride. The organic phase was washed withaqueous KHSO₄(0.2M) and brine, dried and concentrated.

FAB-MS:(m+1)=237(m/z) ¹H-NMR: (400 MHz, CDCl₃) δ 4.82 (s, 2H), 4.22 (s,2H), 3.31-3.26 (t, 2H), 3.10-3.15 (t, 2H), 1.68-1.52 (m, 2H), 1.97-0.86(m, 6H)

(ii) (nPr)₂NC(O)CH₂OOCCH₂—(R)Cgl-Aze-Pab-COO-Bn(4-NO₂)

The title compound was prepared analogously to the procedure describedin Example 1(iii) from H—(R)Cgl-Aze-Pab-COOBn(4-NO₂) (343 mg; 0.62 mmol;see Example 62(ii) above) and (nPr)₂NC(O)CH₂OOCCH₂Cl (160 mg; 0.68 mmol;from step (i) above). Yield 89 mg (19%).

FAB-MS: (m+1)=750(m/z)

EXAMPLE 68 (2-Me)BnOOCCH₂—(R)Cgl-Aze-Pab-COOCH₂OOCtBu

The title compound was prepared analogously to the procedure describedin Example 1(iii) from H—(R)Cgl-Aze-Pab-COOCH₂OOCtBu (380 mg; 0.71 mmol;see Example 12(v) above) and 2-(methyl)benzyl bromoacetate (215 mg; 0.88mmol; see Example 32(i) above). Yield 37 mg (7.5%).

FAB-MS:(m+1)=692(m/z)

EXAMPLE 69

The compounds of Examples 1 to 68 were all tested in Test A above andwere all found to exhibit an IC₅₀TT value of more than 1.0 μM (ie theywere are inactive to thrombin per se; cf. the active inhibitorHOOC—CH₂—(R)Cgl-Aze-Pab-H which exhibits an IC₅₀TT of 0.01 μM).

EXAMPLE 70

The compounds of Examples 1 to 68 were tested in one, two or all ofTests B, C and/or D above, and were all found to exhibit oral and/orparenteral bioavailability in the rat as the active inhibitorHOOC—CH₂—(R)Cgl-Aze-Pab-H, either as the free acid and/or as one or moreester thereof. Based on the assumption that HOOC—CH₂—(R)Cgl-Aze-Pab-H isformed in the rat, the bioavailability was calculated according to theformulae described in Test B and/or Test C as appropriate.

Abbreviations

-   Ac=acetyl-   aq=aqueous-   Aze ═S-Azetidine-2-carboxylic acid-   Boc=t-butyloxycarbonyl-   (Boc)₂O=di-t-butyldicarbonate-   Bn=benzyl-   Bu=butyl-   Cgl=cyclohexylglycine-   Ch=cyclohexyl-   DCC=dicyclohexyl carbodiimide-   DMAP=N,N-dimethyl amino pyridine-   EDC=1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride-   Et=ethyl-   EtOH=ethanol-   EtOAc=ethyl acetate-   h=hours-   HCl=hydrochloric acid-   H-Pab-H=1-amidino-4-aminomethyl benzene-   H-Pab-Z=4-aminomethyl-1-(N-benzyloxycarbonylamidino)benzene-   HPLC=high performance liquid chromatography-   K₂CO₃=anhydrous potassium carbonate-   Me=methyl-   Men=(1R,2S,5R)-menthyl-   Pab-OH=4-aminomethyl-benzamidoxime(4-aminomethyl-1-(amino-hydroxyiminomethyl)benzene-   Piv(aloyl)=2,2-dimethylacetyl-   Pr=propyl-   Prl=N-pyrrolidinyl-   RPLC=reverse phase high performance liquid chromatography-   TFA=trifluoroacetic acid-   THF=tetrahydrofuran-   Z=benzyloxy carbonyl

Prefixes n, s, i and t have their usual meanings: normal, iso, sec andtertiary. Prefixes in the NMR-spectra s, d, t, q, and b mean singlet,doublet, triplet, quartet, and broad, respectively. The stereochemistryfor the amino acids is by default (S) if not otherwise stated.

1. A pharmaceutical formulation comprising an effective amount ofacetylsalicylic acid and a compound of formula I,R¹O(O)C—CH₂—(R)Cgl-Aze-Pab-R²  I wherein R¹ represents —R³ or-A¹(O)N(R⁴)R⁵ in which A¹ represents C₁₋₃ alkylene or -A¹C(O)OR⁴ inwhich A¹ represents C₁₋₅ alkylene; R² (which replaces one of thehydrogen atoms in the amidino unit of Pab-H) represents OH, OC(O)R⁶,C(O)OR⁷ or C(O)OCH(R⁸)OC(O)R⁹; R³ represents H, C₁₋₁₀ alkyl, or C₁₋₃alkylphenyl (which latter group is optionally substituted by C₁₋₆ alkyl,C₁₋₆ alkoxy, nitro or halogen); R⁴ and R⁵ independently represent H,C₁₋₆ alkyl, phenyl, 2-naphthyl or, when R¹ represents -A¹C(O)N(R⁴)R⁵,together with the nitrogen atom to which they are attached representpyrrolidinyl or piperidinyl; R⁶ represents C₁₋₁₇ alkyl, phenyl or2-naphthyl (all of which are optionally substituted by C₁₋₆ alkyl orhalogen); R⁷ represents 2-naphthyl, phenyl, C₁₋₃ alkylphenyl (whichlatter three groups are optionally substituted by C₁₋₆ alkyl, C₁₋₆alkoxy, nitro or halogen), or C₁₋₁₂ alkyl (which latter group isoptionally substituted by C₁₋₆ alkoxy, C₁₋₆ acyloxy or halogen); R⁸represents H or C₁₋₄ alkyl; and R⁹ represents 2-naphthyl, phenyl, C₁₋₆alkoxy or C₁₋₈ alkyl (which latter group is optionally substituted byhalogen, C₁₋₆ alkoxy or C₁₋₆ acyloxy); provided that: when R¹ representsR³, R³ represents benzyl, methyl, ethyl, n-butyl or n-hexyl and R²represents C(O)OR⁷, then R⁷ does not represent benzyl; or apharmaceutically-acceptable salt thereof.
 2. A formulation, as definedin claim 1, wherein, in the compound of formula I, R⁴ represents H orC₁₋₆ alkyl when R¹ represents -A¹C(O)N(R⁴)R⁵.
 3. A formulation, asdefined in claim 1, wherein, in the compound of formula I, R⁵ representsC₁₋₄ alkyl or C₄₋₆ cycloalkyl when R¹ represents -A¹O(O)N(R⁴)R⁵.
 4. Aformulation, as defined in claim 1, wherein, in the compound of formulaI, R⁴ and R⁵ together represent pyrrolidinyl when R¹ represents-A¹C(O)N(R⁴)R⁵.
 5. A formulation, as defined in claim 1, wherein, in thecompound of formula I, A¹ represents C₁₋₃ alkylene, and R⁴ represents Hor C₁₋₃ alkyl and R⁵ represents C₂₋₆ alkyl or C₅₋₆ cycloalkyl, or R⁴ andR⁵ together represent pyrrolidinyl when R¹ represents-A¹C(O)N(R⁴)R⁵. 6.A formulation, as defined in claim 1, wherein, in the compound offormula I, A¹ represents C₁₋₅ alkylene when R¹ represents -A¹C(O)OR⁴. 7.A formulation, as defined in claim 6, wherein A¹ represents C₁₋₅alkylene and R⁴ represents C₁₋₄ alkyl.
 8. A formulation, as defined inclaim 1, wherein, in the compound of formula 1, R⁴ represents C₁₋₆ alkylwhen R¹ represents -A¹C(O)OR⁴.
 9. A formulation, as defined in claim 1,wherein, in the compound of formula I, R³ represents H, C₁₋₁₀ alkyl(which latter group is optionally linear or, when there are a sufficientnumber of carbon atoms, is optionally branched and/or be partiallycyclic or cyclic), or C₁₋₃ alkylphenyl (which latter groups isoptionally substituted, is optionally linear or, when there are asufficient number of carbon atoms, is optionally branched), when R¹represents R³.
 10. A formulation as claimed in claim 1, wherein, in thecompound of formula I, R¹ represents H, linear C₁₋₁₀ alkyl, branchedC₃₋₁₀ alkyl, partially cyclic C₄₋₁₀ alkyl, C₄₋₁₀ cycloalkyl, optionallysubstituted linear C₁₋₃ alkylphenyl, optionally substituted branched C₃alkylphenyl.
 11. A formulation as claimed in claim 10, wherein R¹represents linear C₁₋₆ alkyl, C₆₋₁₀ cycloalkyl, or optionallysubstituted linear C₁₋₃ alkylphenyl.
 12. A formulation, as defined inclaim 1, wherein, in the compound of formula I, R² represents OH.
 13. Aformulation, as defined in claim 1, wherein, in the compound of formulaI, R⁶ represents optionally substituted phenyl or C₁₋₁₇ alkyl (whichlatter group is optionally linear or, when there are a sufficient numberof carbon atoms, is optionally branched, is optionally cyclic orpartially cyclic, and/or is optionally saturated or unsaturated) when R²represents OC(O)R⁶.
 14. A formulation as claimed in claim 13 wherein R⁶represents optionally substituted phenyl, linear C₁₋₄ alkyl, branchedC₁₋₃ alkyl or cis-oleyl.
 15. A formulation as claimed in claim 14wherein R⁶ represents linear C₁₋₃ alkyl or branched C₃ alkyl.
 16. Aformulation, as defined in claim 1, wherein, in the compound of formulaI, R⁷ represents optionally substituted phenyl, C₁₋₁₂ alkyl (whichlatter group is optionally substituted, is optionally linear or, whenthere are a sufficient number of carbon atoms, is optionally branched,cyclic or partially cyclic, and/or saturated or unsaturated), or C₁₋₃alkylphenyl (which latter group is optionally substituted, is optionallylinear or, when there are a sufficient number of carbon atoms, isoptionally branched) when R² represents C(O)OR⁷.
 17. A formulation inclaim 16 wherein R⁷ represents optionally substituted and/or optionallyunsaturated linear C₁₋₄ alkyl or optionally substituted and/oroptionally unsaturated branched C₃₋₄ alkyl, optionally substitutedphenyl, or optionally substituted linear C₁₋₃ alkylphenyl or optionallysubstituted branched C₃ alkylphenyl.
 18. A formulation as claimed inclaim 17 wherein R⁷ represents optionally substituted linear C₁₋₄ alkylor optionally substituted branched C₃₋₄ alkyl, optionally substitutedlinear C₁₋₃ alkylphenyl or branched C₃ alkylphenyl.
 19. A formulation,as defined in claim 1, wherein, in the compound of formula I, R⁸represents H or methyl, when R² represents C(O)OCH(R^(8)OC(O)R) ⁹.
 20. Aformulation, as defined in claim 19 wherein R⁸ represents H or methyland R⁹ represents phenyl, C₅₋₇ cycloalkyl, linear C₁₋₆ alkyl, branchedC₃₋₆ alkyl or partially cyclic C₇₋₈ alkyl.
 21. A formulation as claimedin claim 20 wherein R⁸ represents H and R⁹ represents C₅₋₇ cycloalkyl,linear C₁₋₈ alkyl or partially cyclic C₇₋₈ alkyl.
 22. A formulation, asdefined in claim 1, wherein, in the compound of formula I, R⁹ representsphenyl, or C₁₋₈ alkyl (which latter group is optionally substituted, isoptionally linear or, when there are a sufficient number of carbonatoms, is optionally branched and/or cyclic or partially cyclic) when R²represents C(O)OCH(R⁸)OC(O)R⁹.
 23. A formulation as claimed in claim 1wherein, in the compound of formula I, when R¹ represents R³ and R³represents optionally substituted C₁₋₃ alkylphenyl, the optionalsubstituent C₁₋₆ alkyl.
 24. A formulation as claimed in claim 23 whereinthe substituent is methyl.
 25. A formulation as claimed in claim 1wherein, in the compound of formula I, when R² represents C(O)OR⁷ and R⁷represents optionally substituted C₁₋₁₂ alkyl, the optional substituentis selected from halogen and C₁₋₈ alkoxy.
 26. A formulation as claimedin claim 25 wherein the substituent is selected from chloro and methoxy.27. A formulation as claimed in claim 1 wherein, in the compound offormula I, when R² represents C(O)OR⁷ and R⁷ represents optionallysubstituted phenyl, the optional substituent is selected from C₁₋₆alkyl, C₁₋₆ alkoxy and halogen.
 28. A formulation as claimed in claim 27wherein the substituent is selected from methyl, methoxy and chloro. 29.A formulation as claimed in claim 1 wherein, in the compound of formulaI, when R² represents C(O)OR⁷ and R⁷ represents optionally substitutedC₁₋₃ alkylphenyl, the optional substituent is nitro.
 30. A formulationas claimed in claim 1 wherein the compound of formula I isEtOOCCH₂—(R)Cgl-Aze-Pab-COOCH₂CH═CH₂;nPrOOCCH₂—(R)Cgl-Aze-Pab-COOCH₂CH═CH₂;tBuOOCCH₂—(R)Cgl-Aze-Pab-COOCH₂CH═CH₂; EtOOCCH₂—(R)Cgl-Aze-Pab-COOEt;EtOOCCH₂—(R)Cgl-Aze-Pab-COOnBu; PrlC(O)CH₂CH₂CH₂OOCCH₂—(R)Cgl-Aze-Pa-Z;ChNHC(O)CH₂OOCCH₂—(R)Cgl-Aze-Pab-Z;(nPr)₂NC(O)CH₂OOCCH₂—(R)Cgl-Aze-Pab-COOCH₂OOCC(CH₃)₃;EtOOCCH₂—(R)Cgl-Aze-Pab-COOCH₂OOCC(CH₃);EtOOCCH₂—(R)Cgl-Aze-Pab-COOCH(CH₃)OOCCH₃; MeOOCCH₂—(R)Cgl-Aze-Pab-OOCPh;MeOOCCH₂—(R)Cgl-Aze-Pab-OH; EtOOCCH₂—(R)Cgl-Aze-Pab-OH;BnOOCCH₂—(R)Cgl-Aze-Pab-OH; nPrOOCCH₂—(R)Cgl-Aze-Pab-Z;nPrOOCCH₂—(R)Cgl-Aze-Pab-OH; iPrOOCCH₂—(R)Cgl-Aze-Pab-OH;tBuOOCCH₂—(R)Cgl-Aze-Pab-OH; (nPr)₂NCOCH₂OOCCH₂—(R)Cgl-Aze-Pab-OH;ChNHCOCH₂OOCCH₂—(R)Cgl-Aze-Pab-OH; EtOOCCH₂—(R)Cgl-Aze-Pab-OAc;HOOCCH₂—(R)Cgl-Aze-Pab-OH; HOOCCH₂—(R)Cgl-Aze-Pab-O-cis-Oleyl;Cyclooctyl-OOCCH₂—(R)Cgl-Aze-Pa-Z; tBuCH₂OOCCH₂—(R)Cgl-Aze-Pab-Z;(2-Me)BnOOCCH₂—(R)Cgl-Aze-Pab-Z; ChCH₂OOCCH₂—(R)Cgl-Aze-Pab-Z;ChOOCCH₂—(R)Cgl-Aze-Pab-Z; PhC(Me)₂OOCCH₂—(R)Cgl-Aze-Pab-Z;(Me)₂CHC(Me)₂OOCCH₂—(R)Cgl-Aze-Pab-Z;BnOOCCH₂—(R)Cgl-Aze-Pab-COOPh(4-OMe);ChCH₂OOCCH₂—(R)Cgl-Aze-Pab-COOPh(4-OMe);(2-Me)BnOOCCH₂—(R)Cgl-Aze-Pab-COOPh(4-OMe);EtOOCCH₂—(R)Cgl-Aze-Pab-COOPh(4-Me);BnOOCCH₂—(R)Cgl-Aze-Pab-COOPh(4-Me); BnOOCCH₂—(R)Cgl-Aze-Pab-COO-nBu;iPrOOCCH₂—(R)Cgl-Aze-Pab-COOCH₂CH═CH₂; EtOOCCH₂—(R)Cgl-Aze-Pab-COO-iBu;BnOOCCH₂—(R)Cgl-Aze-Pab-COO-nPr; EtOOCCH₂—(R)Cgl-Aze-Pab-COOCH₂OOCCh;EtOOCCH₂—(R)Cgl-Aze-Pab-COOCH₂OCCH₂Ch;EtOOCCH₂—(R)Cgl-Aze-Pab-COOCH(Me)OOCPh;EtOOCCH₂—(R)Cgl-Aze-Pab-COOCH₂OOCPh;BnOOCCH₂—(R)Cgl-Aze-Pab-COOCH(Me)OAc; EtOOCCH₂—(R)Cgl-Aze-Pab-COOCH₂OAc;tBuOOCCH₂—(R)Cgl-Aze-Pab-COOCH₂OAc;MeOOC—C(═CHEt)CH₂—OOCCH₂—(R)Cgl-Aze-Pab-Z;Men-OOCCH₂—(R)Cgl-Aze-Pab-COOPh(4-OMe); andEtOOCCH₂—(R)Cgl-Aze-Pab-COOCH₂CCl₃.
 31. A formulation as claimed inclaim 1 wherein the compound of formula I isEtOOCCH₂—(R)Cgl-Aze-Pab-COOCH₂CCl₃; BnOOCCH₂—(R)Cgl-Aze-Pab-COOnBu;nPrOOCCH₂—(R)Cgl-Aze-Pab-Z; Cyclooctyl-OOCCH₂—(R)Cgl-Aze-Pab-Z;EtOOCCH₂—(R)Cgl-Aze-Pab-COOCH₂OOCCh; MeOOCCH₂—(R)Cgl-Aze-Pab-OH;EtOOCCH₂—(R)Cgl-Aze-Pab-OH; nPrOOCCH₂—(R)Cgl-Aze-Pab-OH;iPrOOCCH₂—R)Cgl-Aze-Pab-OH; BnOOCCH₂—(R)Cgl-Aze-Pab-OH; andEtOOCCH₂—(R)Cgl-Aze-Pab-OAc.
 32. A formulation, as defined in claim 1,with the additional proviso that, in the compound of formula I, R¹ doesnot represent -A¹C(O)OR⁴.
 33. A formulation, as defined in claim 1, withthe additional proviso that, in the compound of formula I, R⁴ and R⁵ donot independently represent H.
 34. A formulation , as defined in claim1, with the additional proviso that, in the compound of formula I, R⁶does not represent C₁₋₁₇ alkyl, when R² represents OC(O)R⁶.
 35. Aformulation, as defined in claim 1, wherein, in the compound of formulaI, R¹ represents -A¹C(O)OR⁴.
 36. A formulation, as defined in claim 1,wherein, in the compound of formula I, R⁴ and R⁵ independently representH.
 37. A formulation, as defined in claim 1, wherein, in the compound offormula I, R⁶ represents C₁₋₁₇ alkyl, when R² represents OC(O)R⁶.
 38. Aformulation as claimed in claim 1, wherein the compound of formula I isEtOOCCH₂—(R)Cgl-Aze-Pab-OH.
 39. A method of treatment of a conditionwhere inhibition of thrombin is required which method comprisesadministration of a therapeutically effective amount of a formulation asdefined in claim 1 or claim 38, to a person suffering from, orsusceptible to, such a condition for a time and under conditionssuitable for eliciting an antithrombotic effect.
 40. A method as claimedin claim 39, wherein the condition is thrombosis.
 41. A method asclaimed in claim 39, wherein the condition is hypercoagulability inblood and tissues.
 42. A combination product comprising (a)acetylsalicylic acid and (b) a compound of the formula I as defined inclaim
 1. 43. A combination product as claimed in claim 42 whichcomprises a kit of parts comprising components (a) and (b).
 44. Acombination product as claimed in claim 42, in which components (a) and(b) are suitable for sequential, separate and/or simultaneous use in thetreatment of a condition where inhibition of thrombin is required.
 45. Acombination product as claimed in claim 44, wherein component (a) iscombined with component (b).
 46. A method of treatment of a conditionwhere inhibition of thrombin is required, which method comprisesadministration of an effective amount of a combination product asclaimed in claim 42 for a time and under conditions suitable foreliciting an antithrombotic effect.
 47. A method of treatment of acondition where inhibition of thrombin is required, which methodcomprises administration of a therapeutically effective amount ofcomponents (a) and (b) of a combination product as claimed in claim 42separately, sequentially or simultaneously for a time and underconditions suitable for eliciting an antithrombotic effect.
 48. A methodof treatment of thrombosis or of hypercoagulability in blood andtissues, which comprises administration of an effective amount of acombination product as claimed in claim 42 for a time and underconditions suitable for eliciting an antithrombotic effect.
 49. Acombination product comprising (a) acetylsalicylic acid and (b) thecompound EtOOCCH₂—(R)Cgl-Aze-Pab-OH or a pharmaceutically-acceptablesalt thereof.
 50. A method of treatment of a condition where inhibitionof thrombin is required, which method comprises administration of aneffective amount of a combination product as claimed in claim 49 for atime and under conditions suitable for eliciting an antithromboticeffect.
 51. A method of treatment of thrombosis or of hypercoagulabilityin blood and tissues, which comprises administration of an effectiveamount of a combination product as claimed in claim 49 for a time andunder conditions suitable for eliciting an antithrombotic effect.
 52. Acombination product comprising (a) acetylsalicylic acid and (b) thecompound EtOOCCH₂—(R)Cgl-Aze-Pab-OH.
 53. A method of treatment of acondition where inhibition of thrombin is required, which methodcomprises administration of an effective amount of a combination productas claimed in claim 52 for a time and under conditions suitable foreliciting an antithrombotic effect.
 54. A method of treatment ofthrombosis or of hypercoagulability in blood and tissues, whichcomprises administration of an effective amount of a combination productas claimed in claim 52 for a time and under conditions suitable foreliciting an antithrombotic effect.